The first amputated human limb model for studying human tissue imaging probes

The emerging field of fluorescence-guided surgery (FGS) is rapidly developing and has the potential to greatly improve the safety and effectiveness
of surgery.
In FGS, tissues of interest are targeted and special molecular markers
called fluorophores are used.
The primary function of these fluorophores is to distinguish the tissue of interest from other tissues and subsequently guide surgical steps
.
Currently, FDA approval for fluorophores for clinical use is limited to three: indocyanine green (ICG), fluorescein, and methylene blue (MB).

While these drugs have several clinical applications, they are non-targeted, which limits their specificity
.
The growing demand for FGS has spurred the identification of new, specific fluorophores that target specific tissues and are expected for clinical success
.
Although many candidate fluorophores have been shown to be effective in animal models, their clinical translation requires rigorous testing and significant financial investment
.

A recent study published in the Journal of Biomedical Optics (JBO) addresses this question
.
In this study, researchers from the United States developed a new, improved system for identifying fluorescent agents
with the highest chance of clinical success.
Logan M.
Bateman, lead author of the study, said, "Understanding how these fluorophores behave in human tissues is critical
to improving the accuracy and safety of fluorescent agents and ultimately reducing development costs and minimizing potential harm to patients.
"

To reduce the time for fluorophore selection, the researchers collaborated
with Gibbs' lab at Oregon Health & Science University.
Together, they developed an amputated human lower limb model for testing nerve-specific fluorophores
.
In this model, tissue examination begins shortly after amputation, prior to
tissue breakdown caused by hypoxia.
Using a heart-pumped saline injection, the team was able to simulate blood vessels and osmotic pressure
in living human tissues.

After amputation, the limbs are transported to a surgical laboratory, where targeted fluorophores
are perfused using a circulatory loop.
First, normal saline is perfused through the main artery, followed by a standard dose of LGW16-03 fluorophore (neurospecific).

The limbs are drained by gravity, and the collected perfusion fluid is circulated back to the loop, simulating blood circulation
.
This occurs during 10 min fluorophore perfusion, followed by 20 min
rinse with normal saline.
After 30 min, the neural tissue is imaged in situ (inside the amputated limb) and in vitro (detached from the limb) using open-field and closed-field fluorescence imaging systems
.

Given that the agent is non-toxic, the key parameter that determines its effectiveness is its signal-background ratio (SBR), which is an indicator
of the desired signal (i.
e.
, signal from neural tissue) relative to background noise.
Notably, fluorophores exhibit excellent performance
in this regard.

"We are impressed with the SBR using this fluorophore and believe it will also perform clinically well
.
" By looking at these contrasts, we are confident that the perfusion model adequately delivers fluorophores to the tissue of interest," commented senior author Eric R.
Henderson at Dartmouth-Hitchcock Medical Center
.
The results confirmed that the formulation had the optical properties required to highlight the target tissue and, more importantly, demonstrated the feasibility
of this new model of amputated human limbs.

So, what does the future hold? The team believes that human limb models can in the future be used not only to study and select other fluorescent agents, but also to study peripheral diseases and pathological features
of tissues under controlled conditions.
In addition, the team applied the platform to study changes
caused by tumor growth.

Henderson concludes: "This is the first in vitro model to examine the performance of this fluorophore, and while further testing is needed, the model has great potential
for many different applications in translation research.
"