Juntendo Medical Journal

Atherosclerotic cardiovascular disease is the leading cause of death worldwide. Among the most promising results of the preventing approach against progression of atherosclerosis derived from lipid lowering therapy especially by statins. Recent clinical trials with statins indicated that aggressive lipid lowering therapy could inhibit the progression of atherosclerosis or even induce the regression of atherosclerotic plaque. It also stabilizes the unstable plaque to stable form and reduces clinical CV events. Several trials using intravascular ultrasound demonstrated that lowering LDL-C to 70-80 mg/dl by aggressive lipid-lowering treatment with moderately high-dose statins significantly reduced coronary plaque volume, but these studies are vastly underpowered to determine the effects of such lipid-lowering treatment on clinical outcomes. Accordingly, the value of reducing LDL-C levels substantially below 100 mg/dl in patients with CAD, particularly Asian patients, had not been clearly demonstrated, and Japanese guidelines recommended reducing LDL-C only to immediately below 100 mg/dl until recently. HIJ-PROPER and REAL-CAD provided pivotal evidence for the aggressive lipid-lowering in Japanese patients at secondary prevention. The lipid management strategy for secondary prevention is still changing in clinical practice from recent Japanese evidence. In this article, we will review the history and current understanding of the efficacy of aggressive lipid lowering therapy which inhibits the atherosclerosis progression and induces the regression, and we also will review the recent clinical evidence especially from Japan.


Introduction
Muscle atrophy is involved in chronic heart failure 1) . We have previously revealed that angiotensin II (Ang II), a key mediator of the reninangiotensin system, directly induced muscle atrophy in mice via activation of NADPH oxidase (Nox), a major source of reactive oxygen species (ROS) 2) . In this study, we assessed the role of Nox4 and insulin-like growth factor 1 receptor (IGF-1R) in muscle atrophy in Ang II-infusion mice.

Materials and Methods
Twelve-week old male Nox4 knockout (KO) mice and age-matched male wild-type (WT) mice were used. All experimental and animal care procedures were approved by our institutional animal research committee and conformed to the guidelines for the Care and Use of Laboratory Animals at Juntendo University Graduate School of Medicine. Either saline (vehicle) or Ang II (1,000 ng/kg/min) was infused into WT and Nox4 KO mice via subcutaneously implanted osmotic minipumps for 4 weeks. Experiments were performed in following 4 groups; WT + V, Nox4 KO + V, WT + Ang II, and Nox4 KO + Ang II. Lower skeletal muscle limbs were removed and used for Nox activity, quantitative real-time PCR and western blot analysis.

Results
At baseline, there were no differences in physical characteristics between WT and Nox4 KO mice. At 4 weeks after Ang II-infusion, WT + Ang II mice showed an increase of NADPH oxidase activation in the skeletal muscle, which was inhibited in Nox4 KO + Ang II mice. Body and lower skeletal muscle weight were significantly decreased in WT + Ang II mice compared with WT + V mice. These changes were significantly restored in Nox4 KO + Ang II mice. Protein expression level of Akt phosphorylation of Ser473, key molecule of protein synthesis, was decreased in WT + Ang II mice compared with WT + V mice. This parameter was significantly restored in Nox4 KO + Ang II mice (all p-values < 0.05). However, gene expression level of IGF-1R was no differences in all groups (Figure-1).

Discussion
The aims of this study were to determine the effect of Nox4 deficiency on Ang II-induced muscle atrophy in mice. For this purpose, we used Nox4 KO and WT mice.
Insulin signaling, including IGF-1R and Akt controls vital growth, survival and metabolic functions in the skeletal muscle and other tissues 3) . Therefore, we evaluated the Akt phosphorylation and IGF-1R in the skeletal muscle. In the present study, Nox4 deficiency attenuates AII-induced muscle atrophy. Moreover, consistent with this result, decrease in Akt phosphorylation at Ser473 was restored in Nox4 KO mice. However, gene expression level of IGF-1R was no differences in all groups. Brink et al. have been reported that circulating IGF-I levels, IGF-binding protein (IGFBP)-3, and IGFBP-5 were significant reduced in the skeletal muscle of Ang II-infusion rats 4) . Thus, IGF-1 and IGFBPs needs to evaluate in the skeletal muscle and plasma in our future study.

Conclusion
Angiotensin II induces muscle atrophy via NADPH oxidase 4-derived ROS production, suggesting that Nox4 plays an important role in the development of muscle atrophy.