calpain and Peritonitis

Calpains are ubiquitous pro-inflammatory proteases, whose activity is controlled by calpastatin, their specific inhibitor. Transgenic mice over-expressing rabbit calpastatin (CalpTG) are protected against vascular remodelling and angiotensin II-dependent inflammation. We hypothesized that specific calpain inhibition would protect against aging-related lesions in arteries and kidneys. We analysed tissues from 2-months and 2-years-old CalpTG and wild-type mice and performed high throughput RNA-Sequencing of kidney tissue in aged mice. In addition, we analysed inflammatory response in the kidney of aged CalpTG and wild-type mice, and in both in vivo (monosodium urate peritonitis) and in vitro models of inflammation. At two years, CalpTG mice had preserved kidney tissue, less vascular remodelling and less markers of senescence than wild-type mice. Nevertheless, CalpTG mice lifespan was not extended, due to the development of lethal spleen tumors. Inflammatory pathways were less expressed in aged CalpTG mice, especially cytokines related to NF-κB and NLRP3 inflammasome activation. CalpTG mice had reduced macrophage infiltration with aging and CalpTG mice produced less IL-1α and IL-1β in vivo in response to inflammasome activators. In vitro, macrophages from CalpTG mice produced less IL-1α in response to particulate activators of inflammasome. Calpains inhibition protects against inflammaging, limiting kidney and vascular lesions related to aging.

Topics: Aging; Animals; Arteries; Calcium-Binding Proteins; Calpain; Cells, Cultured; Cysteine Proteinase Inhibitors; Cytokines; Inflammasomes; Kidney; Mice; Mice, Inbred C57BL; NF-kappa B; NLR Family, Pyrin Domain-Containing 3 Protein; Peritonitis; Rabbits

Activation of the innate immune system is critical for clearance of bacterial pathogens to limit systemic infections and host tissue damage. Here, we report a key role for calpain proteases in bacterial clearance in mice with acute peritonitis. Using transgenic mice expressing Cre recombinase primarily in innate immune cells (fes-Cre), we generated conditional capns1 knockout mice. Consistent with capns1 being essential for stability and function of the ubiquitous calpains (calpain-1, calpain-2), peritoneal cells from these mice had reduced levels of calpain-2/capns1, and reduced proteolysis of their substrate selenoprotein K. Using an acute bacterial peritonitis model, we observed impaired bacterial killing within the peritoneum and development of bacteremia in calpain knockout mice. These defects correlated with significant reductions in IL-1α release, neutrophil recruitment, and generation of reactive oxygen species in calpain knockout mice with acute bacterial peritonitis. Peritoneal macrophages from calpain knockout mice infected with enterobacteria ex vivo, were competent in phagocytosis of bacteria, but showed impaired clearance of intracellular bacteria compared with control macrophages. Together, these results implicate calpains as key mediators of effective innate immune responses to acute bacterial infections, to prevent systemic dissemination of bacteria that can lead to sepsis.

Topics: Acute Disease; Animals; Bacterial Infections; Calpain; Disease Models, Animal; Interleukin-1alpha; Macrophages, Peritoneal; Mice; Mice, Knockout; Neutrophil Infiltration; Peritonitis; Phagocytosis; Reactive Oxygen Species

Zymosan enhances the formation of reactive oxygen species, which contributes to the pathophysiology of multiple organ failure. We investigated the effects of calpain inhibitor I (5, 10, or 20 mg/kg) on the multiple organ failure caused by zymosan (500 mg/kg, administered intraperitoneally as a suspension in saline) in rats.. University research laboratory.. Male Sprague-Dawley rats.INTERVENTIONS Multiple organ failure in rats was assessed 18 hrs after administration of zymosan and/or calpain inhibitor I and was monitored for 12 days (for loss of body weight and mortality rate).. Treatment of rats with calpain inhibitor I (5, 10, or 20 mg/kg intraperitoneally, 1 and 6 hrs after zymosan) attenuated the peritoneal exudation and the migration of polymorphonuclear cells caused by zymosan in a dose-dependent fashion. Calpain inhibitor I also attenuated the lung, liver, and intestinal injury (histology) as well as the increase in myeloperoxidase activity and malondialdehyde concentrations caused by zymosan in the lung, liver, and intestine. Immunohistochemical analysis for nitrotyrosine and for poly(adenosine-disphosphate-ribose) revealed positive staining in lung, liver, and intestine from zymosan-treated rats. The degree of staining for nitrotyrosine and poly(adenosine-disphosphate-ribose) was reduced markedly in tissue sections obtained from zymosan-treated rats administered calpain inhibitor I (20 mg/kg intraperitoneally). Furthermore, treatment of rats with calpain inhibitor I significantly reduced the expression of inducible nitric oxide synthase and cyclooxygenase-2 in lung, liver, and intestine.. This study provides the first evidence that calpain inhibitor I attenuates the degree of zymosan-induced multiple organ failure in the rat.

Topics: Animals; Calpain; Cell Movement; Cyclooxygenase 2; Dose-Response Relationship, Drug; Glycoproteins; Immunohistochemistry; Intestinal Mucosa; Intestines; Isoenzymes; Liver; Lung; Male; Malondialdehyde; Multiple Organ Failure; Neutrophils; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Peritoneum; Peritonitis; Peroxidase; Peroxynitrous Acid; Poly Adenosine Diphosphate Ribose; Prostaglandin-Endoperoxide Synthases; Rats; Rats, Sprague-Dawley; Tyrosine; Zymosan