Establishment of a system for screening autophagic flux regulators using a modified fluorescent reporter and CRISPR/Cas9.

Autophagy is a mechanism of bulk protein degradation that plays an important role in regulating homeostasis in many organisms. Among several methods for evaluating its activity, a fluorescent reporter GFP-LC3-RFP-LC3ΔG, in which GFP-LC3 is cleaved by ATG4 following autophagic induction and degraded in lysosome, has been used for monitoring autophagic flux, which is the amount of lysosomal protein degradation. In this study, we modified this reporter by exchanging GFP for pHluorin, which is more sensitive to low pH, and RFP to mCherry, to construct pHluorin-LC3-mCherry reporter.

In-Depth Insight Into the Mechanisms of Cardiac Dysfunction in Patients With Type 1 Diabetes Mellitus Using Layer-Specific Strain Analysis.

Background: Although the subclinical left ventricular (LV) dysfunction caused by diabetes mellitus (DM) results in a high risk of death and heart failure, the details of cardiac dysfunction across a wide age range remain unclear. The aim of this study was to assess LV dysfunction in patients with type 1 DM (T1DM) using layer-specific strain analysis by echocardiography.

Methods and results: The 52 patients (median age: 23 [range: 5-40] years) with T1DM were divided into 3 age groups (D1: 5-14 years, D2: 15-24 years, D3: 25-40 years); 78 age- and sex-similar controls were divided into 3 corresponding groups (C1, C2, and C3).