A fluorogenic near-infrared imaging agent for quantifying plasma and local tissue renin activity in vivo and ex vivo.

The renin-angiotensin system (RAS) is well studied for its regulation of blood pressure and fluid homeostasis, as well as for increased activity associated with a variety of diseases and conditions, including cardiovascular disease, diabetes, and kidney disease. The enzyme renin cleaves angiotensinogen to form angiotensin I (ANG I), which is further cleaved by angiotensin-converting enzyme to produce ANG II. Although ANG II is the main effector molecule of the RAS, renin is the rate-limiting enzyme, thus playing a pivotal role in regulating RAS activity in hypertension and organ injury processes. Our objective was to develop a near-infrared fluorescent (NIRF) renin-imaging agent for noninvasive in vivo detection of renin activity as a measure of tissue RAS and in vitro plasma renin activity. We synthesized a renin-activatable agent, ReninSense 680 FAST (ReninSense), using a NIRF-quenched substrate derived from angiotensinogen that is cleaved specifically by purified mouse and rat renin enzymes to generate a fluorescent signal. This agent was assessed in vitro, in vivo, and ex vivo to detect and quantify increases in plasma and kidney renin activity in sodium-sensitive inbred C57BL/6 mice maintained on a low dietary sodium and diuretic regimen. Noninvasive in vivo fluorescence molecular tomographic imaging of the ReninSense signal in the kidney detected increased renin activity in the kidneys of hyperreninemic C57BL/6 mice. The agent also effectively detected renin activity in ex vivo kidneys, kidney tissue sections, and plasma samples. This approach could provide a new tool for assessing disorders linked to altered tissue and plasma renin activity and to monitor the efficacy of therapeutic treatments.

Noninvasive in vivo quantification of neutrophil elastase activity in acute experimental mouse lung injury.

We developed a neutrophil elastase-specific near-infrared fluorescence imaging agent, which, combined with fluorescence molecular tomographic imaging, allowed us to detect and quantify neutrophil elastase activity in vivo, in real time, and noninvasively in an acute model of lung injury (ALI). Significantly higher fluorescent signal was quantified in mice with LPS/fMLP-induced ALI as compared to healthy controls, correlating with increases in the number of bronchoalveolar lavage cells, neutrophils, and elastase activity. The agent was significantly activated ex vivo in lung sections from ALI but not from control mice, and this activation was ablated by the specific inhibitor sivelestat. Treatment with the specific inhibitor sivelestat significantly reduced lung signal in mice with ALI. These results underscore the unique ability of fluorescence molecular imaging to quantify specific molecular processes in vivo, crucial for understanding the mechanisms underlying disease progression and for assessing and monitoring novel pharmacological interventions.