ABSTRACT
Acute pancreatitis is a disease associated with inflammation and tissue damage. One protein that protects against acute injury, including ischemic injury to both the kidney and heart, is renalase, which is secreted into the blood by the kidney and other tissues. However, whether renalase reduces acute injury associated with pancreatitis is unknown. Here, we used both in vitro and in vivo murine models of acute pancreatitis to study renalase's effects on this condition. In isolated pancreatic lobules, pretreatment with recombinant human renalase (rRNLS) blocked zymogen activation caused by cerulein, carbachol, and a bile acid. Renalase also blocked cerulein-induced cell injury and histological changes. In the in vivo cerulein model of pancreatitis, genetic deletion of renalase resulted in more severe disease, and administering rRNLS to cerulein-exposed WT mice after pancreatitis onset was protective. Because pathological increases in acinar cell cytosolic calcium levels are central to the initiation of acute pancreatitis, we also investigated whether rRNLS could function through its binding protein, plasma membrane calcium ATPase 4b (PMCA4b), which excretes calcium from cells. We found that PMCA4b is expressed in both murine and human acinar cells and that a PMCA4b-selective inhibitor worsens pancreatitis-induced injury and blocks the protective effects of rRNLS. These findings suggest that renalase is a protective plasma protein that reduces acinar cell injury through a plasma membrane calcium ATPase. Because exogenous rRNLS reduces the severity of acute pancreatitis, it has potential as a therapeutic agent.
Subject(s)
Monoamine Oxidase/metabolism , Pancreas/metabolism , Pancreatitis/metabolism , Plasma Membrane Calcium-Transporting ATPases/metabolism , Acinar Cells/drug effects , Acinar Cells/metabolism , Acinar Cells/pathology , Animals , Anti-Inflammatory Agents, Non-Steroidal/metabolism , Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Biomarkers/metabolism , Calcium Signaling/drug effects , Carbachol/pharmacology , Cell Line , Ceruletide/toxicity , Enzyme Activation/drug effects , Fluorescent Antibody Technique, Indirect , Gene Expression Regulation, Enzymologic/drug effects , Humans , Hypertension/etiology , Hypertension/prevention & control , Ligands , Membrane Transport Modulators/pharmacology , Mice , Mice, Knockout , Monoamine Oxidase/blood , Monoamine Oxidase/genetics , Monoamine Oxidase/therapeutic use , Pancreas/drug effects , Pancreas/immunology , Pancreas/pathology , Pancreatitis/chemically induced , Pancreatitis/drug therapy , Pancreatitis/pathology , Plasma Membrane Calcium-Transporting ATPases/antagonists & inhibitors , Plasma Membrane Calcium-Transporting ATPases/chemistry , Plasma Membrane Calcium-Transporting ATPases/genetics , Recombinant Fusion Proteins/metabolism , Recombinant Fusion Proteins/therapeutic use , Taurolithocholic Acid/analogs & derivatives , Taurolithocholic Acid/pharmacologyABSTRACT
Neutron crystallography is a powerful technique for experimental visualization of the positions of light atoms, including hydrogen and its isotope deuterium. In recent years, structural biologists have shown increasing interest in the technique as it uniquely complements X-ray crystallographic data by revealing the positions of D atoms in macromolecules. With this regained interest, access to macromolecular neutron crystallography beamlines is becoming a limiting step. In this report, it is shown that a rapid data-collection strategy can be a valuable alternative to longer data-collection times in appropriate cases. Comparison of perdeuterated rubredoxin structures refined against neutron data sets collected over hours and up to 5 d shows that rapid neutron data collection in just 14 h is sufficient to provide the positions of 269 D atoms without ambiguity.
