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Elemental imaging of human tissue: clinical therapy support and development of new diagnostics

Periodic Reporting for period 1 - TissueMaps (Elemental imaging of human tissue: clinical therapy support and development of new diagnostics)

Reporting period: 2018-06-01 to 2020-05-31

TissueMaps is as a multidisciplinary project of Jožef Stefan Institute (JSI) and University Medical Centre Maribor (UKC MB) with the aim of improving the diagnostic techniques available at hospitals to study prosthesis failures.

Improved living conditions, better health systems and medical innovation have contributed to increased life expectancy in developed countries. Among medical treatments, hip replacements have become a widely used solution. Worldwide, approximately one million hip replacements take place every year representing a financial burden on health systems.

Degradation of hip prostheses results in the release of debris and metallic particles into the surrounding tissue, possibly causing inflammation and prosthesis loosening. Despite hip prosthesis complications due to degradation are relatively frequent, there is no complete understanding of the processes driving this degradation.

Techniques currently applied in hospitals, such as X-ray imaging, magnetic resonance imaging (MRI) or optical tissue microscopies are not able to provide the chemical picture (i.e. content of elements such as Titanium, Aluminium, Vanadium or Zirconium released by the prosthesis) of the surrounding tissue. We use a broad spectrum of advanced microscopy techniques to map elements released by prosthesis into the body, among which Proton-Induced X-ray Emission with focused high-energy proton beam (micro-PIXE) is the main technique.

This project incorporates novel cutting-edge diagnostic tools to study the complex mechanism of dispersion of metallic particles from the prosthesis into the human tissue and the potential link with prosthesis failures and complications.

So far, the development and testing procedures of hip prosthesis have been carried out almost exclusively by the orthopaedic industry. Our work will allow for independent research in this field contributing to a better understanding of the processes involved in prosthesis failures and rejections.
"Tissue samples surrounding the faulty prosthesis are obtained during replacement surgery, by biopsy, at UKC MB. We have studied 12 cases, of which 7 males and 5 females, representing different cases of prosthesis failure.

PIXE analysis allows us to “see” things that we cannot see with “ordinary” optical microscopy. This technique helps us to identify the elements present in the tissue by recording their fingerprints, obtained from the emitted X-rays. How do we do that? PIXE is based on the excitation of an atom, ionized after the passage of a fast particle (proton), and the measurement of the energy released when an electron “jumps” to fill the hole (Figure 1). This decay energy is released as X-ray emission, characteristic of each element (its fingerprint).
The analysis is carried out at the 2 MV tandem accelerator, at JSI, in Ljubljana (Slovenia). Tissue samples are placed inside a vacuum chamber, and scanned with a 3 MeV focused proton beam. The X-ray emitted are collected by two detectors: Silicon Drift Detector and intrinsic Germanium (SDD and iGe).

Figure 2 (a) presents, as example, the X-ray image taken during the preparation for replacement surgery from Patient 2, showing a modular neck fracture, a common problem of this type of bimodular endoprostheses. On Figure 2 (b), a picture of the tissue slice is shown together with the Titanium PIXE map. The elemental map highlights that the surrounding tissue accumulated Titanium debris from degraded prosthesis.

Micro-PIXE has allowed determining the content, size and nature of debris found in the tissue. Oxidation state of the elements originated from the prosthesis have been studied with X-ray Absorption Spectroscopy (XAS) at Synchrotron of Elettra (Italy). Fluorescence Lifetime Imaging Microscopy (FLIM) helped identify fluorophores and nanoparticles in the tissue, and to correlate the increased autofluorescence signal of fluorophores in presence of metallic particles. This had never been studied until today.

The dissemination of results included the ER attendance and active participation in 5 Scientific Conferences, 5 Trainings and 3 Workshops. She presented the results of the research, connected with other researchers, and explored collaboration opportunities with other research groups.

The project results were posted in an open access article, websites and social media (links below):
- Open access article: https://doi.org/10.1016/j.nimb.2019.10.019
- Article available at ZENODO repository: https://zenodo.org/record/3689496#.X2M5qGgzaUk
- Project’s site: https://tissuemaps.wordpress.com/
- JSI Nature publication coverage on Slovenian TV, mention to the prosthesis study: https://www.ionbeamcenters.eu/radiate/dissemination/video-clips/
- JSI, Department of Low and Medium Energy Physics: https://f2.ijs.si/en
- Fellow’s personal LinkedIn, Instagram and Facebook accounts."
Techniques applied at Hospital have been complemented with the introduction of novel analytical methodologies and techniques that enable a detailed study of the body response to implants, by mapping the elemental composition of affected tissue.

During the routine examination at Hospital, X-ray scanning can identify when a prosthesis has failed, pathological examination can give a more detailed information by analysing a tissue sample obtained by biopsy. However, these techniques cannot identify the content, size and origin of debris found in the body. This is crucial to let doctors gain better understanding of hip prosthesis failures.

Hip prostheses and medical devices can be made of different materials depending on the device requirements, manufacturing process and cost considerations. Hip prostheses heads are usually ceramic, while the rest might be titanium alloy. The information about elements in tissue given by PIXE can help identify which type of components are more prone to cause prosthesis degradation and complications, and which type create less complication.

Results of this research may have an impact on the prosthesis industry, by improving the tests before marketing a new prosthesis, or phasing out those prosthesis showing high rates of metallic release.

Therefore, this research may have positive implications to the health systems and on people’s quality of life by improving the diagnostic techniques.

The project has offered a unique possibility to conduct independent research on hip prosthesis, given that it has not been sponsored or co-financed by any prosthesis manufacturer.

The success of TissueMaps project has been possible thanks to a close cooperation between JSI and UKC MB. Interest on this topic is on the rise, and we have started new collaborations with Hospitals from Switzerland and Germany.
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Figure 1