Autophagy and Age-Related Pathologies

Autophagy is a regulated process of self-cannibalization which helps cells maintain homeostasis and regain energy [1,2]. Under basal conditions, autophagy can be used for such things as the degradation of long-lived proteins, but it is mainly induced in response to stress. Under stress like an infection, it is known to target pathogens for lysosomal degradation. During metabolic stress, which is common for cancer cells, autophagy provides ATP for cellular activity and preserves cell viability. Cancer cells induce autophagy where the cells eat themselves and by doing so also protect themselves from cell death. Autophagy is characterized by the formation of autophagosomes and involves several steps: initiation, nucleation, elongation, maturation and degradation. While autophagy is linked to health, the relationship between autophagy and age-related pathologies, such as neurodegenerative diseases, remains unclear [2]. A better understanding of that relationship may lead to the development of clinical applications that promote long-term health.

The nematode, C. elegans, is a well-studied model organism which provides an opportunity for investigating mechanisms of autophagy and age-related pathologies in the context of a whole organism [3,4]​​​​​​​.

How authophagy and age-related diseases can be studied in more detail with a THUNDER Imager are demonstrated in this article.

When imaging C. elegans worms, a solution that can quickly achieve sharp, high-contrast 3D images, where important details are clearly resolved, is most practical. Conventional widefield microscopy is fast and offers detection sensitivity, but unfortunately images of thick specimens, like whole organisms, often show an out-of-focus blur or haze which reduces the contrast [5]​​​​​​​.

Roudworms, C. elegans, were used for this study. The worms were expressing MAH215 [6], GFP, and mCherry. MAH215 is a dual-fluorescent mCherry:GFP:LGG-1 protein which can monitor autophagic flux by visualizing both autophagosomes as well as autolysosomes. GFP (green) indicates autophagosomes and mCherry (red) the autolysosomes that quench GFP in the acidic environment resulting in the emission of the mCherry signal. The worms were imaged with a THUNDER Imager Model Organism. Small volume computational clearing (SVCC) [5]​​​​​​​​​​​​​​ was applied to the image data set and a maximum intensity projection was generated.

The THUNDER Imager Model Organism enables sharp stereoscopic and macroscopic imaging of C. elegans worms by the removal of the out-of-focus light when compared to conventional widefield microscopy [5]​​​​​​​. It allows cell processes to be studied and quantified in more detail.

The THUNDER technology Small Volume Computational Clearing (SVCC) [5] significantly enhances the contrast when imaging C. elegans worms, enabling highly detailed and sharper stereoscopic images to be resolved compared to conventional widefield microscopy. The superior imaging capability provided by the THUNDER technology may lead to a better understanding of the relationship between autophagy and age-related diseases.

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