Polyglutamine diseases are inherited neurodegenerative diseases characterized by misfolding and aggregation of proteins possessing expanded polyglutamine repeats. As overexpression of some heat shock protein (Hsp) suppresses polyglutamine aggregates and cell death, it is assumed that combined overexpression of Hsps will suppress that more effectively. Here, we examined the impact of active forms of heat shock transcription factor 1 (HSF1), which induces a set of Hsps, on polyglutamine inclusion formation and disease progression. We found that active HSF1 suppressed polyglutamine inclusion formation more significantly than any combination of Hsps in culture cells, possibly by regulating expression of unknown genes, as well as major Hsps. We crossed R6/2 Huntington disease mice with transgenic mice expressing an active HSF1 (HSF1Tg). Analysis of the skeletal muscle revealed that the polyglutamine inclusion formation and its weight loss were improved in R6/2/ HSF1Tg mice. Unexpectedly, the life span of R6/2/HSF1Tg mice was significantly improved, although active HSF1 is not expressed in the brain. These results indicated that active HSF1 has a strong inhibitory effect on polyglutamine aggregate formation in vivo and in vitro.

t has been thought that lysosomes carry the non-selective bulk of protein degradation that occurs in cellular remodeling and the removal of abnormal cellular components. 1 In the process of lysosomal degradation, lysosomal membranes and lysosomal enzymes play important roles. Lysosome-associated membrane proteins are thought to be structural or functional components of the lysosomal membrane. 2 Cathepsins are hydrolytic enzymes in lysosomes that degrade damaged proteins. Lysosomal function is broadly categorized into autophagy and heterophagy. Autophagy is the process of sequestration of intracellular components and their subsequent degradation by the lysosomal vacuoles. In contrast, sequestered cell organelles derived exogenously from other cells are degraded in the lysosomes through a process called heterophagy. 3 It is reported that, under the conditions of sublethal injury such as ischemia, cardiomyocytes contain autophagic vacuoles ranging from those in which organelles are readily identified to those characteristic of residual bodies. 4,5 Morphological studies of cardiomyopathic hearts have revealed degenerative changes such as vacuolization and myofibrillar lysis. 6 In hearts from cases of dilated cardiomyopathy Japanese Circulation Journal Vol. 65, November 2001 (DCM), increased activity of lysosomal enzymes is found, 7-9 but the precise mechanism of lysosomal function in the pathogenesis of DCM remains poorly understood. It is speculated that autophagic function is associated with the irreversible degeneration preceding myocardial cell death. MethodsTwenty-seven patients with idiopathic DCM (20 men, 7 women; mean age, 47±14 years) who had undergone partial left ventriculectomy were included in the study. 10,11 Specimens of left ventricular myocardium, resected during surgery, were examined and for the control, 5 autopsied hearts from patients who had died of non-cardiac diseases were used. Light MicroscopyThe formalin-fixed tissues were embedded in paraffin. Sections were cut, deparafinized, stained with hematoxylineosin and observed under a light microscope. ImmunohistochemistryIn situ localization of cathepsin D, a lysosomal enzyme, and lysosome-associated membrane protein-1 (LAMP-1) were examined immunohistochemically, using an anticathepsin D antibody (Upstate biotechnology, NY, USA) and an anti-LAMP-1 antibody (Santa Cruz, CA, USA), respectively. The sections were incubated with the biotinylated secondary antibody and the avidin-biotin peroxidase complex method was carried out. Positive reactions were optically detected by 3,3'-diaminobenzidine. Sections were counterstained with hematoxylin and examined under a light microscope. Autophagic Degeneration as a Possible Mechanism of Myocardial Cell Death in Dilated CardiomyopathyHiroaki Shimomura, MD; Fumio Terasaki, MD; Tetsuya Hayashi, MD; Yasushi Kitaura, MD; Tadashi Isomura, MD*; Hisayoshi Suma, MD*In failing hearts, cardiomyocytes degenerate and interstitial fibrosis, which indicates cardiomyocyte loss, becomes more prominent in the myocardium...

Objective-Vascular endothelial growth factor (VEGF) plays an important role in inducing angiogenesis. Mesenchymal stem cells (MSCs) may have potential for differentiation to several types of cells, including myocytes. We hypothesized that transplantation of VEGF-expressing MSCs could effectively treat acute myocardial infarction (MI) by providing enhanced cardioprotection, followed by angiogenic effects in salvaging ischemic myocardium. Methods and Results-The human VEGF 165 gene was transfected to cultured MSCs of Lewis rats using an adenoviral vector. Six million VEGF-transfected and LacZ-transfected MSCs (VEGF group), LacZ-transfected MSCs (control group), or serum-free medium only (medium group) were injected into syngeneic rat hearts 1 hour after left coronary artery occlusion. At 1 week after MI, MSCs were detected by X-gal staining in infarcted region. High expression of VEGF was immunostained in the VEGF group. At 28 days after MI, infarct size, left ventricular dimensions, ejection fraction, E wave/A wave ratio and capillary density of the infarcted region were most improved in the VEGF group, compared with the medium group. Immunohistochemically, ␣-smooth muscle actin-positive cells were most increased in the VEGF group. Key Words: angiogenesis Ⅲ gene therapy Ⅲ myocardial infarction Ⅲ stem cell Ⅲ transplantation C ell transplantation has become a promising novel therapy for ischemic heart disease and heart failure. Recent studies have revealed that various types of cells are effective in cell transplantation after myocardial infarction (MI), such as skeletal myoblasts, 1,2 smooth muscle cells, 3 and bone marrow mononuclear cells. 4 Bone marrow mononuclear cells are especially useful because they contain, among various lineage cells, hematopoietic cells and endothelial progenitor cells; therefore they have the ability to induce angiogenesis in ischemic tissue. A reported clinical trial of cell transplantation with skeletal myoblasts and mononuclear bone marrow cells showed that such therapies can have cardioprotective and angiogenic effects after MI. 5,6 However, selection of the most appropriate cell types for transplantation is controversial. Conclusions-ThisMesenchymal stem cells (MSCs) are isolated from bone marrow mononuclear cells and can be expanded ex vivo. Under appropriate culture conditions, MSCs have the potential to terminally differentiate into osteocytes, chondrocytes, adipocytes, tenocytes, myotubes, astrocytes, hematopoietic supporting stroma, and endothelial cells. 7 MSCs have also been used in a model of cell transplantation, 8,9 showing that these cells could differentiate into myogenic cells. Therefore, MSCs have many characteristics that make them useful for cellular therapy.Vascular endothelial growth factor (VEGF) is a strong therapeutic reagent for treating ischemia by inducing angiogenesis. 10 It has been reported that direct intramyocardial gene transfer results in localized enhancement of VEGF levels and successful angiogenesis in animal models of MI. 11 Furthermore, recent h...

Abnormalities in Z-disc proteins cause hypertrophic (HCM), dilated (DCM) and/or restrictive cardiomyopathy (RCM), but disease-causing mechanisms are not fully understood. Myopalladin (MYPN) is a Z-disc protein expressed in striated muscle and functions as a structural, signaling and gene expression regulating molecule in response to muscle stress. MYPN was genetically screened in 900 patients with HCM, DCM and RCM, and disease-causing mechanisms were investigated using comparative immunohistochemical analysis of the patient myocardium and neonatal rat cardiomyocytes expressing mutant MYPN. Cardiac-restricted transgenic (Tg) mice were generated and protein-protein interactions were evaluated. Two nonsense and 13 missense MYPN variants were identified in subjects with DCM, HCM and RCM with the average cardiomyopathy prevalence of 1.66%. Functional studies were performed on two variants (Q529X and Y20C) associated with variable clinical phenotypes. Humans carrying the Y20C-MYPN variant developed HCM or DCM, whereas Q529X-MYPN was found in familial RCM. Disturbed myofibrillogenesis with disruption of α-actinin2, desmin and cardiac ankyrin repeat protein (CARP) was evident in rat cardiomyocytes expressing MYPN(Q529X). Cardiac-restricted MYPN(Y20C) Tg mice developed HCM and disrupted intercalated discs, with disturbed expression of desmin, desmoplakin, connexin43 and vinculin being evident. Failed nuclear translocation and reduced binding of Y20C-MYPN to CARP were demonstrated using in vitro and in vivo systems. MYPN mutations cause various forms of cardiomyopathy via different protein-protein interactions. Q529X-MYPN causes RCM via disturbed myofibrillogenesis, whereas Y20C-MYPN perturbs MYPN nuclear shuttling and leads to abnormal assembly of terminal Z-disc within the cardiac transitional junction and intercalated disc.