Results 8 MPEG-PL-Cy5.5 Probe Characterization In order to determine trypsin activation in experimental pancreatitis using the caerulein model, we developed a probe that was selective to trypsin activity. To evaluate the mechanism of action, in vivo, of trypsin inhibitor drug-like compounds, we prepared a self-quenched selleck chemicals llc trypsin activatable near infrared smart probe. When rat anionic trypsin was added to 1 ��M of mPEG-PL-Cy5.5 probe, increase in fluorescence intensity was observed which was positively correlated to the concentration of trypsin enzyme and the time of incubation. To establish trypsin selectivity, mPEG-PL-Cy5.5 probe activation was tested by addition to a panel of endopeptidases that are described to be expressed in the pancreas, including human and rat pancreatic elastase, kallikrein 1, kallikrein 5, chymotrypsin, cathepsin L, and cathepsin B [22].

As shown in figure 1, mPEG-PL-Cy5.5 probe was activated in the presence of trypsin in contrast to other pancreatic enzymes. In the presence of the highly specific trypsin inhibiting protein SPINK1, the protease activation was suppressed to background levels (figure 1). Therefore, it can be concluded that mPEG-PL-Cy5.5 probe was highly trypsin selective. Figure 1 MPEG-PL-Cy5.5 probe characterization. 9 In Vivo Monitoring of Trypsin Activation 9.1 Model development The caerulein-injection model is a well-established mechanistic model for experimental pancreatitis. For optical imaging, however, the anatomical location of the pancreas posed a challenge to signal acquisition.

Animal weight and age were critical when choosing subjects for optical imaging study. Adult rats were considered the most suitable because of their smaller stomach size and separation of the pancreas and stomach. Upon administration of caerulein in rats, edema development is observed. We administered a blood pool imaging agent Angiosense 680 to confirm our ability to image pancreas non-invasively in rats. Healthy animals that were subjected to repeated doses of caerulein showed increasing accumulation of the blood pool agent Angiosense 680. The graph in figure 2a shows a threefold enhancement in Angiosense 680 signal in the pancreas normalized to signal acquired before caerulein administration. Therefore, for these studies, three caerulein administrations were sufficient. Figure 2 In vivo pancreatitis imaging of edema (A) and trypsin activation (B).

Establishing the ability to image diseased pancreas in a rat model using optical imaging, we evaluated the time course and severity of trypsin activation in this experimental pancreatitis model using the trypsin activatable mPEG-PL-Cy5.5 probe. 9.2 Trypsin dependent activation of mPEG-PL-Cy5.5 probe MPEG-PL-Cy5.5 probe was administered to healthy animals and then three hourly repeated caerulein injections were GSK-3 administered to evaluate the time and severity of trypsin activation. MPEG-PL-Cy5.