2. Insightful Analysis
Here, a molecular imaging protocol served as a platform to test the capacity of 99m
Tc-GA-5 (a radiolabeled anti-GFAP) to identify in vivo the astrocytic response in the spinal cord of rats subjected to SCI with a model that reproduces human SCI 
To accomplish this goal, before performing the imaging studies, we made sure that the radiolabeling efficiency and the radiotracer integrity met the required standards. We also confirmed, by radioimmunohistochemistry, that the RIC is bound to the cells and tissues of interest in a post-injury time-dependent manner. After solving these basic precepts, we designed the imaging protocol grounded on SPECT utilizing a dedicated small animal system (Albira small animal microPET/SPECT/CT imaging system). As a global result, our radiotracer selectively accumulates in glial cells (targeting primarily reactive astrocytes) as an active agent, facilitating the in vivo monitoring of astrogliosis.
GA-5′s receptors can be over or underexpressed in cells and tissues under physiological or pathological conditions and used as molecular targets 
. The specific interaction between the receptor and its ligand (GFAP and 99m
Tc-anti-GFAP in our study) proved to be an effective strategy to evaluate this radiopharmaceutical’s amount and residence time in target tissues. By considering the temporal/spatial expression of GFAP in the current radioimmunohistochemistry assay (see Figure 1
) and previous histological studies 
, we chose to perform our imaging study at 20 days after injury, as this is the timeframe where reactive astrocytes overexpress GFAP in the context of the glial scar formation.
Figure 1. %Activity in post-injury histological samples (n = 3–6) on different days, illustrating the affinity of 99mTc-GA-5 for its receptors in cells of murine medullary tissue.
Contrary to most in vivo molecular imaging studies where the radioactive biomarker is administered intravenously, we used the intrathecal route successfully. When macromolecules, such as proteins, are administered intravenously, the blood-brain barrier limits their access to the CNS. However, when delivered intrathecally, macromolecules circumvent this barrier and effectively reach their target within the CNS 
. The imaging strategy tested here was effective in revealing the cellular and molecular processes sought. The biomarker appears to be highly sensitive to GFAP expression changes and the distribution of the reactive astrocytes at and around the injury site.
SCI is a pathology with high physical, social, and economic repercussions 
commonly occurring in vehicular accidents, sports-related injuries, or violent episodes, for which no therapies to restore neurologic deficit exist effectively. After an injury occurs, death of neurons and glial cells, ischemia, and inflammation, which is followed by the formation of a glial scar and cystic cavities in the spinal cord, are observed 
. Due to the profound impact of astrogliosis on the progression of SCI, a better understanding of the cellular and molecular events in glial scarring is mandatory 
. To date, the role of astrogliosis has been oversimplified in binary terms as good or bad 
. It is considered beneficial because it aids in repairing the initial damage, stabilizes the spread of injury, and fosters axonal regeneration and functional recovery, but detrimental because it provides both a physical and chemical barrier to regenerating axons 
There is no diagnostic strategy for in vivo selective identification of GFAP present in reactive astrocytes through molecular imaging methods. Here, we demonstrate that the rapid identification of astrogliosis through molecular imaging in a murine SCI model is feasible without characterizing the mechanisms that regulate astrocyte reactivity and scar formation. We consider that the main contribution of this study lies in the development of a tool that will be useful to understand better and monitor astrogliosis, a controversial topic of the most significant relevance in the pathophysiology of SCI.
Nuclear imaging (SPECT and PET scans) is widely used in cardiology, neurology, and oncology. Although no studies have been reported evaluating molecular imaging technologies targeting GFAP in SCI, some recent studies have reported the utility of nuclear imaging to reveal in vivo biological activity of events associated with SCI, such as acute inflammation in a rodent model