Aims The mitochondria are important sources of reactive oxygen species (ROS) in the heart. Mitochondrial ROS production has been implicated in the pathogenesis of diabetic cardiomyopathy, suggesting that therapeutic strategies specifically targeting mitochondrial ROS may have benefit in this disease. We investigated the therapeutic effects of mitochondria-targeted antioxidant mito-TEMPO on diabetic cardiomyopathy. Methods The mitochondria-targeted antioxidant mito-TEMPO was administrated after diabetes onset in a mouse model of streptozotocin-induced type-1 diabetes and type-2 diabetic db/db mice. Cardiac adverse changes were analyzed and myocardial function assessed. Cultured adult cardiomyocytes were stimulated with high glucose, and mitochondrial superoxide generation and cell death were measured. Results Incubation with high glucose increased mitochondria superoxide generation in cultured cardiomyocytes, which was prevented by mito-TEMPO. Co-incubation with mito-TEMPO abrogated high glucose-induced cell death. Mitochondrial ROS generation, and intracellular oxidative stress levels were induced in both type-1 and type-2 diabetic mouse hearts. Daily injection of mito-TEMPO for 30 days inhibited mitochondrial ROS generation, prevented intracellular oxidative stress levels, decreased apoptosis and reduced myocardial hypertrophy in diabetic hearts, leading to improvement of myocardial function in both type-1 and type-2 diabetic mice. Incubation with mito-TEMPO or inhibition of Nox2-containing NADPH oxidase prevented oxidative stress levels and cell death in high glucose-stimulated cardiomyocytes. Mechanistic study revealed that the protective effects of mito-TEMPO were associated with down-regulation of ERK1/2 phosphorylation. Conclusions Therapeutic inhibition of mitochondrial ROS by mito-TEMPO reduced adverse cardiac changes and mitigated myocardial dysfunction in diabetic mice. Thus, mitochondria-targeted antioxidants may be an effective therapy for diabetic cardiac complications.

Aims/hypothesis MicroRNAs (miRs) have been suggested as potential therapeutic targets for heart diseases. Inhibition of miR-195 prevents apoptosis in cardiomyocytes stimulated with palmitate and transgenic overexpression of miR-195 induces cardiac hypertrophy and heart failure. We investigated whether silencing of miR-195 reduces diabetic cardiomyopathy in a mouse model of streptozotocin (STZ)-induced type 1 diabetes. Methods Type 1 diabetes was induced in C57BL/6 mice (male, 2 months old) by injections of STZ. Results MiR-195 expression was increased and levels of its target proteins (B cell leukaemia/lymphoma 2 and sirtuin 1) were decreased in STZ-induced type 1 and db/db type 2 diabetic mouse hearts. Systemically delivering an anti-miR-195 construct knocked down miR-195 expression in the heart, reduced caspase-3 activity, decreased oxidative stress, attenuated myocardial hypertrophy and improved myocardial function in STZ-induced mice with a concurrent upregulation of B cell leukaemia/lymphoma 2 and sirtuin 1. Diabetes reduced myocardial capillary density and decreased maximal coronary blood flow in mice. Knockdown of miR-195 increased myocardial capillary density and improved maximal coronary blood flow in diabetic mice. Upregulation of miR-195 sufficiently induced apoptosis in cardiomyocytes and attenuated the angiogenesis of cardiac endothelial cells in vitro. Furthermore, inhibition of miR-195 prevented apoptosis in cardiac endothelial cells in response to NEFA, an important feature of diabetes. Conclusions/interpretation Therapeutic silencing of miR-195 reduces myocardial hypertrophy and improves coronary blood flow and myocardial function in diabetes, at least in part by reducing oxidative damage, inhibiting apoptosis and promoting angiogenesis. Thus, miR-195 may represent an alternative therapeutic target for diabetic heart diseases.

Liver metastasis is a clinically significant contributor to the mortality associated with melanoma, colon, and breast cancer. Preclinical mouse models are essential to the study of liver metastasis, yet their utility has been limited by the inability to study this dynamic process in a noninvasive and longitudinal manner. This study shows that three-dimensional high-frequency ultrasound can be used to noninvasively track the growth of liver metastases and evaluate potential chemotherapeutics in experimental liver metastasis models. Liver metastases produced by mesenteric vein injection of B16F1 (murine melanoma), PAP2 (murine H-ras-transformed fibroblast), HT-29 (human colon carcinoma), and MDA-MB-435/HAL (human breast carcinoma) cells were identified and tracked longitudinally. Tumor size and location were verified by histologic evaluation. Tumor volumes were calculated from the three-dimensional volumetric data, with individual liver metastases showing exponential growth. The importance of volumetric imaging to reduce uncertainty in tumor volume measurement was shown by comparing threedimensional segmented volumes with volumes estimated from diameter measurements and the assumption of an ellipsoid shape. The utility of high-frequency ultrasound imaging in the evaluation of therapeutic interventions was established with a doxorubicin treatment trial. These results show that three-dimensional high-frequency ultrasound imaging may be particularly well suited for the quantitative assessment of metastatic progression and the evaluation of chemotherapeutics in preclinical liver metastasis models.