Glaucoma is a chronic degenerative disease that affects the optic nerve and is the leading cause of irreversible blindness in the world. During the course of the disease, there is a progressive loss of retinal ganglion cells and their axons (cells that make up the optic nerve) with consequent anatomical alterations (excavation of the head of the optic nerve and loss of retinal nerve fibers). The functional consequence is represented by a progressive narrowing of the visual field which, in the absence of treatment, can lead to the complete loss of visual function. In Italy it is estimated that almost one million people suffer from glaucoma, and that about half of them are unaware of it.

Glaucoma, in fact, runs apparently asymptomatic for a long time and, often, vision problems occur suddenly. Unfortunately, when the first symptoms appear, the damage is already irreversible. Blindness caused by glaucoma can be prevented, if the disease is diagnosed and treated promptly. Although several factors have been identified that increase the risk of glaucoma, the cause of the disease remains unknown.

Key risk factors include increased eye pressure, familiarity, thin corneal thickness, and age. The major health problems related to glaucoma are linked to the need to efficiently identify affected individuals in the population, to the need to guarantee specialist visits and periodic diagnostic examinations to the identified individuals and to the development of increasingly accurate technologies in making increasingly earlier diagnosis of the disease.

From a scientific point of view, the main fields of interest concern the understanding of the mechanisms underlying the onset and progression of damage, the search for disease biomarkers, the development of technologies that allow a certain and early diagnosis and the development of new therapeutic strategies both medical and surgical.

Proteomics Laboratory

The research activity of the proteomics laboratory is focused on the development and application of mass spectrometry protocols for the study of biological samples isolated from patients belonging to the IRCCS Bietti Foundation, according to the procedures indicated in the study protocols approved by the local Ethics Committees.

Overall, the scientific activity is aimed at:

• Understand the pathophysiological bases of ocular diseases, through the search for proteome alterations in the biological samples under examination;
• Set up and carry out biomarker discovery programmes for the diagnosis and follow-up of eye diseases.

In order to carry out these ambitious research programs, the laboratory has progressively equipped itself with state-of-the-art instrumentation, such as: Orbitrap Exploris 240 in line with a nanoHPLC system (Ultimate3000). In addition, a FAIMS module for mass spectrometry studies in ion-mobility mode was also acquired. The Orbitrap system supplied, in addition to having the latest generation technical requirements, boasts the possibility of setting up and carrying out innovative approaches such as single-cell proteomics.

In support of the research activity in the proteomics field, both for the management of the pre-analytical and sample preparation phase, and for the confirmation of the data obtained in proteomics through different approaches, the laboratory is further equipped with the following instrumentation (the list is not to be considered complete):

• SpeedVac system, for the concentration of samples under atmospheric vacuum conditions;
• analytical centrifugation;
• varioskan lux spectrofluorimeter;
• gel documentation system for visible development, chemiluminescence and fluorescence of gels and filters;
• electrophoresis apparatus complete with Trans-Blot system for ultra-fast dry transfer on nitrocellulose filters.

In order to maximize the scientific growth of the laboratory, as well as its visibility at national and international level, several research agreements have been stipulated with institutions, such as the Universities of Rome Tor Vergata and Roma Tre, the University of Catania, the University of Turin. They are also active collaborations with different international institutes including the University of British Columbia (Vancouver, Canada), the University of Paris Cergy (Paris, France).

The first year of the laboratory's research activity was dedicated to the design and preparation of mass spectrometry analysis protocols with particular attention to shot-gun (through trypsin digestion) and top-down (analysis of proteins under native conditions) approaches, as well as the testing and commissioning of all the previously indicated instrumentation.

However, during 2023, some important scientific objectives were already achieved, which made it possible to achieve (at the turn of the following year) the writing of 6 scientific publications.

In particular, the first proteome characterization of circulating immune cells isolated from glaucoma patients (and controls) was completed.

General research objectives

Glaucoma research, through collaboration between clinic and laboratory, has the ultimate goal of preserving the visual function and quality of life of glaucoma patients:

1. assessment of the diagnostic accuracy and clinical utility of advanced morphological diagnostic tools of the optic nerve head applied in the various stages of glaucomatous pathology
2. Evaluation of the influence of corneal biomechanical properties on the accuracy of applanation tonometry, on the progression of glaucoma and on the efficacy of common ocular hypotensive pharmacological agents under normal conditions and after refractive surgery
3. Study of the epidemiology of glaucomatous disease in the population of the city of Rome
4. Study of the role of inflammatory proteins in the pathogenesis of primary open-angle glaucoma
5. Evaluation of the traditional perimeter and new perimeter strategies in the early diagnosis and study of the progression of eye diseases
6. Evaluation of pharmacological and non-pharmacological therapeutic strategies in the treatment of glaucoma and ocular hypertension