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In vivo imaging with fluorescent smart probes to assess treatment strategies for acute pancreatitis.

Agarwal A, Boettcher A, Kneuer R, Sari-Sarraf F, Donovan A, Woelcke J, Simic O, Brandl T, Krucker T - PLoS ONE (2013)

Bottom Line: A dose dependent decrease of total pancreatic fluorescence signal occurred upon administration of known trypsin inhibitors.The fluorescence-based method was a better predictor of trypsin inhibition than pancreatic to body weight ratio.This method is more sensitive and dynamic than classic tissue sample readouts and could be applied to preclinically optimize trypsin inhibitors towards intrapancreatic target inhibition.

View Article: PubMed Central - PubMed

Affiliation: Novartis Institute of BioMedical Research, Cambridge, Massachusetts, USA.

ABSTRACT

Background and aims: Endoprotease activation is a key step in acute pancreatitis and early inhibition of these enzymes may protect from organ damage. In vivo models commonly used to evaluate protease inhibitors require animal sacrifice and therefore limit the assessment of dynamic processes. Here, we established a non-invasive fluorescence imaging-based biomarker assay to assess real-time protease inhibition and disease progression in a preclinical model of experimental pancreatitis.

Methods: Edema development and trypsin activation were imaged in a rat caerulein-injection pancreatitis model. A fluorescent "smart" probe, selectively activated by trypsin, was synthesized by labeling with Cy5.5 of a pegylated poly-L-lysine copolymer. Following injection of the probe, trypsin activation was monitored in the presence or absence of inhibitors by in vivo and ex vivo imaging.

Results: We established the trypsin-selectivity of the fluorescent probe in vitro using a panel of endopeptidases and specific inhibitor. In vivo, the probe accumulated in the liver and a region attributed to the pancreas by necropsy. A dose dependent decrease of total pancreatic fluorescence signal occurred upon administration of known trypsin inhibitors. The fluorescence-based method was a better predictor of trypsin inhibition than pancreatic to body weight ratio.

Conclusions: We established a fluorescence imaging assay to access trypsin inhibition in real-time in vivo. This method is more sensitive and dynamic than classic tissue sample readouts and could be applied to preclinically optimize trypsin inhibitors towards intrapancreatic target inhibition.

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Related in: MedlinePlus

In vivo pancreatitis imaging of edema (A) and trypsin activation (B).A). Animals (n = 3) were imaged before and after administering the blood pool agent Angiosense 680. Animals were then administered three subsequent doses of caerulein and imaged at various time points. B). Animals (n = 3) were administered the mPEG-PL-Cy5.5 probe 1 h before first caerulein injection. Data represented mean ± SEM.
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pone-0055959-g002: In vivo pancreatitis imaging of edema (A) and trypsin activation (B).A). Animals (n = 3) were imaged before and after administering the blood pool agent Angiosense 680. Animals were then administered three subsequent doses of caerulein and imaged at various time points. B). Animals (n = 3) were administered the mPEG-PL-Cy5.5 probe 1 h before first caerulein injection. Data represented mean ± SEM.

Mentions: 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.


In vivo imaging with fluorescent smart probes to assess treatment strategies for acute pancreatitis.

Agarwal A, Boettcher A, Kneuer R, Sari-Sarraf F, Donovan A, Woelcke J, Simic O, Brandl T, Krucker T - PLoS ONE (2013)

In vivo pancreatitis imaging of edema (A) and trypsin activation (B).A). Animals (n = 3) were imaged before and after administering the blood pool agent Angiosense 680. Animals were then administered three subsequent doses of caerulein and imaged at various time points. B). Animals (n = 3) were administered the mPEG-PL-Cy5.5 probe 1 h before first caerulein injection. Data represented mean ± SEM.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3569412&req=5

pone-0055959-g002: In vivo pancreatitis imaging of edema (A) and trypsin activation (B).A). Animals (n = 3) were imaged before and after administering the blood pool agent Angiosense 680. Animals were then administered three subsequent doses of caerulein and imaged at various time points. B). Animals (n = 3) were administered the mPEG-PL-Cy5.5 probe 1 h before first caerulein injection. Data represented mean ± SEM.
Mentions: 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.

Bottom Line: A dose dependent decrease of total pancreatic fluorescence signal occurred upon administration of known trypsin inhibitors.The fluorescence-based method was a better predictor of trypsin inhibition than pancreatic to body weight ratio.This method is more sensitive and dynamic than classic tissue sample readouts and could be applied to preclinically optimize trypsin inhibitors towards intrapancreatic target inhibition.

View Article: PubMed Central - PubMed

Affiliation: Novartis Institute of BioMedical Research, Cambridge, Massachusetts, USA.

ABSTRACT

Background and aims: Endoprotease activation is a key step in acute pancreatitis and early inhibition of these enzymes may protect from organ damage. In vivo models commonly used to evaluate protease inhibitors require animal sacrifice and therefore limit the assessment of dynamic processes. Here, we established a non-invasive fluorescence imaging-based biomarker assay to assess real-time protease inhibition and disease progression in a preclinical model of experimental pancreatitis.

Methods: Edema development and trypsin activation were imaged in a rat caerulein-injection pancreatitis model. A fluorescent "smart" probe, selectively activated by trypsin, was synthesized by labeling with Cy5.5 of a pegylated poly-L-lysine copolymer. Following injection of the probe, trypsin activation was monitored in the presence or absence of inhibitors by in vivo and ex vivo imaging.

Results: We established the trypsin-selectivity of the fluorescent probe in vitro using a panel of endopeptidases and specific inhibitor. In vivo, the probe accumulated in the liver and a region attributed to the pancreas by necropsy. A dose dependent decrease of total pancreatic fluorescence signal occurred upon administration of known trypsin inhibitors. The fluorescence-based method was a better predictor of trypsin inhibition than pancreatic to body weight ratio.

Conclusions: We established a fluorescence imaging assay to access trypsin inhibition in real-time in vivo. This method is more sensitive and dynamic than classic tissue sample readouts and could be applied to preclinically optimize trypsin inhibitors towards intrapancreatic target inhibition.

Show MeSH
Related in: MedlinePlus