Calpains mediate acute renal cell death: role of autolysis and translocation (original) (raw)
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Calpains Mediate Calcium and Chloride Influx During the Late Phase of Cell Injury
Journal of Pharmacology and Experimental Therapeutics, 1997
The role of Ca ϩϩ in cell death is controversial. Extracellular Ca ϩϩ influx and calpain activation occurred during the late phase of renal proximal tubule cell injury produced by the mitochondrial inhibitor antimycin A. Chelation of intracellular Ca ϩϩ , extracellular Ca ϩϩ , the calcium channel blocker nifedipine, calpain inhibitor 1 and the dissimilar calpain inhibitor PD150606 blocked antimycin A-induced influx of extracellular Ca ϩϩ and cell death. The calcium channel blocker verapamil was ineffective. Calpain inhibitor 1 and PD150606 were cytoprotective also against tetrafluoroethyl-L-cysteine-, bromohydroquinone-, oxidant (t-butylhydroperoxide)-and calcium ionophore (ionomycin)-induced cell death. Extracellular Ca ϩϩ influx was associated with the translocation of calpain activity from the cytosol to the membrane and was prevented by calpain
Endoplasmic reticulum Ca2+ signaling and calpains mediate renal cell death
Cell Death & Differentiation, 2002
The goal of the current study was to determine the roles of ATP content, endoplasmic reticulum (ER) Ca 2+ stores, cytosolic free Ca 2+ (Ca 2+ f) and calpain activity in the signaling of rabbit renal proximal tubular (RPT) cell death (oncosis). Increasing concentrations (0.3 ± 10 mM) of the mitochondrial inhibitor antimycin A produced rapid ATP depletion that correlated to a rapid and sustained increase in Ca 2+ f , but not phospholipase C activation. The ER Ca 2+-ATPase inhibitors thapsigargin (5 mM) or cyclopiazonic acid (100 mM) alone produced similar but transient increases in Ca 2+ f. Pretreatment with thapsigargin prevented antimycin A-induced increases in Ca 2+ f and antimycin A pretreatment prevented thapsigargin-induced increases in Ca 2+ f. Calpain activity increased in conjunction with ER Ca 2+ release. Pretreatment, but not post-treatment, with thapsigargin or cyclopiazonic acid prevented antimycin A-induced cell death. These data demonstrate that extensive ATP depletion signals oncosis through ER Ca 2+ release, a sustained increase in Ca 2+ f and calpain activation. Depletion of ER Ca 2+ stores prior to toxicant exposure prevents increases in Ca 2+ f and oncosis.
Toxicological Sciences, 2006
Calpains and endoplasmic reticulum (ER) stress have both been implicated in renal cell death following exposure to reactive chemical toxicants (RCTs). Therefore, we explored the link between ER stress, calpain, and cell death in renal cell injury due to model RCTs (iodoacetamide, menadione, tert-butyl hydroperoxide) and ER stress inducers (tunicamycin [TUN], thapsigargin [THAPS]). The calpain inhibitor, PD150606, significantly reduced the RCT and TUN-induced cell death in the renal cell line LLC-PK1, but not death induced by THAPS. ER stress was confirmed by the significant induction of GRP78 following exposure to RCTs and ER stress inducers. While GRP94 induction was observed following RCTs and TUN, it was not statistically significant because of variability. THAPS at 5mM significantly induced GRP94, while 20mM caused a calpain-dependent cleavage of GRP94. Caspase-12 and m-calpain were variably induced and/ or cleaved following exposure to all toxicants, supporting activation of these signaling pathways. Inhibition of calpain blocked the induction of GRP78 following exposure to RCTs suggesting that calpain was contributing to the observed ER stress following RCTs. In contrast, calpain inhibition did not block ER stress protein induction following exposure to nontoxic concentrations of TUN or THAPS, indicating that calpain inhibition did not block the ER stress protein induction pathways directly. These studies demonstrate a previously unappreciated link between calpain activation and ER stress-associated cell death in renal cells. While further studies are required to clarify the molecular events involved, these results confirm that calpain activation and the ER are important related players in chemically induced renal cell damage.
Calpains, mitochondria, and apoptosis
Cardiovascular Research, 2012
Mitochondrial activity is critical for efficient function of the cardiovascular system. In response to cardiovascular injury, mitochondrial dysfunction occurs and can lead to apoptosis and necrosis. Calpains are a 15-member family of Ca 2+-activated cysteine proteases localized to the cytosol and mitochondria, and several have been shown to regulate apoptosis and necrosis. For example, in endothelial cells, Ca 2+ overload causes mitochondrial calpain 1 cleavage of the Na + /Ca 2+ exchanger leading to mitochondrial Ca 2+ accumulation. Also, activated calpain 1 cleaves Bid, inducing cytochrome c release and apoptosis. In renal cells, calpains 1 and 2 promote apoptosis and necrosis by cleaving cytoskeletal proteins, which increases plasma membrane permeability and cleavage of caspases. Calpain 10 cleaves electron transport chain proteins, causing decreased mitochondrial respiration and excessive activation, or inhibition of calpain 10 activity induces mitochondrial dysfunction and apoptosis. In cardiomyocytes, calpain 1 activates caspase 3 and poly-ADP ribose polymerase during tumour necrosis factor-a-induced apoptosis, and calpain 1 cleaves apoptosis-inducing factor after Ca 2+ overload. Many of these observations have been elucidated with calpain inhibitors, but most calpain inhibitors are not specific for calpains or a specific calpain family member, creating more questions. The following review will discuss how calpains affect mitochondrial function and apoptosis within the cardiovascular system.
Kidney International, 2003
Calpain is activated in experimental uremia: Is calpain a media-Uremia has long been associated with a high incidence tor of uremia-induced myocardial injury? of cardiovascular mortality [1], but little is known of the Background. The cysteine proteases calpain and caspase-3 mechanism of how uremia causes myocardial injury. The are known mediators of cell death. The aim of this study was aim of this study was to determine whether the cytosolic to assess their contribution to the tissue damage found in experimental uremia. cysteine proteases calpain and caspase-3 have a role in Methods. Calpain and caspase-3 activities were measured in the development of uremia-induced tissue injury. Both the hearts of rats that were sham-operated (control), shamproteases have many common substrates and both cleave operated and spontaneously hypertensive (SHR), and those cytoskeleton-associated proteins that have a role in the rendered uremic by 5/6 nephrectomy (uremic). In an in vitro study, heart myoblasts (Girardi) were incubated with human structural integrity of the cell membrane [2]. Calpains serum from healthy subjects (control serum conditioned media, are present in all mammalian tissues and at least 14 CSCM) or uremic patients (uremic serum conditioned media, isoforms of calpain have been identified [3]. Calpain-1 USCM), in the presence and absence of calpain and caspase-3 (-calpain) and calpain-2 (m-calpain) are the best charinhibitors. After 48 hours the activity of calpain and caspase-3 was measured, and cell injury determined by DNA fragmentaacterized isoforms and both are known to be present tion (ELISA) and lactate dehydrogenase (LDH) release. An in cardiac tissue, with calpain-2 being predominant [4]. in situ assay was designed to study how USCM affects calpain Caspase-3 is considered to be a common downstream activity over time. apoptosis effector, while calpain has been implicated Results. In the in vivo study, mean calpain activities were almost identical in the control and SHR groups, but calpain in both apoptotic and necrotic pathways [2]. Calpain is and caspase-3 activities were much elevated in the uremic group known to be a mediator of rat hypoxic/ischemic and (P Ͻ 0.01 and 0.001 respectively vs. control). The SHR group ischemic/reperfusion injury in the myocardium [5, 6], had significantly higher mean arterial blood pressure (P Ͻ and of multiple organ injury/dysfunction in hemorrhagic 0.001 vs. control, 0.01 vs. uremic). In the in vitro study calpain activity and DNA fragmentation were markedly higher in shock/reperfusion injury [7]. In addition, calpain has re-USCM treated cells compared to CSCM (both PϽ0.05). Both cently been implicated as an intermediary of acute renal were reduced in USCM cells containing calpain inhibitors (E64d, cell death in a hypoxia/reoxygenation model [8]. Calcium calpastatin, or PD 150606). LDH release was raised also in is a prerequisite for calpain activation and is known to USCM treated cultures (P Ͻ 0.05), which only the E64d treatment could significantly reduce (P Ͻ 0.02). Caspase-3 activities be elevated subsequent to ischemia/reperfusion injury were similar in USCM and CSCM groups. The in situ assay [9, 10]. Caspase-3 activity is also heightened in myocarshowed significant increases in calpain activity in USCM dial ischemia [11], and non-selective caspase inhibition treated cells compared to CSCM after just 3.5 hours (PϽ0.01). has been shown to reduce myocardial infarct size in an Conclusions. In vivo results suggest that the increases in calpain and caspase-3 activity in uremic rat hearts were primarily ischemic/reperfusion rat model [12]. due to uremia and not to hypertension. In vitro data demon-We postulated that calpain and/or caspase-3 are actistrate that uremia-induced cell injury can be attenuated by calvated in the uremic heart and therefore might participate pain inhibition. Therefore, it is likely that calpain is a mediator in the development of uremia-induced myocardial injury. of uremia-induced myocardial injury. The role of calpain seems particularly relevant, as calcium homeostasis is known to be altered in the uremic
Inhibitors of calpain activation (PD150606 and E-64) and renal ischemia-reperfusion injury
Biochemical Pharmacology, 2005
Calpain activation has been implicated in the development of ischemia-reperfusion (I-R) injury. Here we investigate the effects of two inhibitors of calpain activity, PD150606 and E-64, on the renal dysfunction and injury caused by I-R of rat kidneys in vivo. Male Wistar rats were administered PD150606 or E-64 (3 mg/kg i.p.) or vehicle (10%, v/v, DMSO) 30 min prior to I-R. Rats were subjected to bilateral renal ischemia (45 min) followed by reperfusion (6 h). Serum and urinary biochemical indicators of renal dysfunction and injury were measured; serum creatinine (for glomerular dysfunction), fractional excretion of Na + (FE Na , for tubular dysfunction) and urinary N-acetylb-D-glucosaminidase (NAG, for tubular injury). Additionally, kidney tissues were used for histological analysis of renal injury, immunohistochemical analysis of intercellular adhesion molecule-1 (ICAM-1) expression and nitrotyrosine formation. Renal myeloperoxidase (MPO) activity (for polymorphonuclear leukocyte infiltration) and malondialdehyde (MDA) levels (for tissue lipid peroxidation) were determined. Both PD150606 and E-64 significantly reduced the increases in serum creatinine, FE Na and NAG caused by renal I-R, indicating attenuation of renal dysfunction and injury and reduced histological evidence of renal damage caused by I-R. Both PD150606 and E-64 markedly reduced the evidence of oxidative stress (ICAM-1 expression, MPO activity, MDA levels) and nitrosative stress (nitrotyrosine formation) in rat kidneys subjected to I-R. These findings provide the first evidence that calpain inhibitors can reduce the renal dysfunction and injury caused by I-R of the kidney and may be useful in enhancing the tolerance of the kidney against renal injury associated with aortovascular surgery or renal transplantation. #
Calpains: physiological and pathophysiological significance
Pathophysiology, 1999
The system of Ca-dependent neutral proteases, commonly known as the calpain system, is represented by two main isoforms, calpain 1 and calpain 2, as well as by tissue-specific enzymes. These enzymes take part in some physiological processes and are responsible for certain pathological states. The versatility of intracellular Ca 2 + mediating the majority of cellular processes is reflected on the calpain system, i.e. it activates the functional reserves of the cell at moderate Ca 2 + concentrations and causes protein degradation at excessive Ca 2 + . Besides, the activity of the calpain system of the cell is regulated by the concentration of its specific inhibitor, calpastatin, as well as by phosphorylation-dephosphorylation of proteins. Calpains can function both inside the cell and outside it (in the extracellular matrix) and exist both in soluble and membrane-bound states, where it is the most active. Calpains induce cell degradation in some pathologies and in apoptosis. Preliminary modification of proteins or their membrane microenvironment initiates enhanced proteolysis of cytosolic and membrane-bound proteins and enzymes. The key role of calpains in the development of many pathological states testifies to the high pathophysiological significance of this proteolytic system in the organism.
Mitochondrial calpain system: An overview
Calpain system is generally known to be comprised of three molecules: two Ca 2+ -dependent proteases: land m-calpains, and their endogenous inhibitor, calpastatin. While calpains have previously been considered as the cytoplasmic enzymes, research in the recent past demonstrated that l-calpain, m-calpain and calpain 10 are present in mitochondria, which play important roles in a variety of pathophysiological conditions including necrotic and apoptotic cell death phenomena. Although a number of original research articles on mitochondrial calpain system are available, yet to the best of our knowledge, a precise review article on mitochondrial calpain system has, however, not been available. This review outlines the key features of the mitochondrial calpain system, and its roles in several cellular and biochemical events under normal and some pathophysiological conditions.
Nephrology, 1996
Acute renal failure (ARF) is a clinical syndrome that is frequently present in hospitalized patients and is associated with high mortality. The duration of the clinical course of ARF correlates with a mortality that remains high despite the availability of dialysis treatment. The virtual complete recovery of renal function in those patients who survive ARF, as well as the minimal renal histological abnormalities, suggest that there are reversible components in the pathophysiology of ARF. A better understanding of the pathogenesis of ARF is therefore needed to allow interventions that would prevent the need for haemodialysis and improve survival. A role of cytoplasmic free calcium [CaZ+], in hypoxia-induced proximal tubule damage has been proposed. To further investigate the role of [Ca2+], in mediating hypoxic proximal tubular injury, a video imaging technique has been developed in which [Caz+Ii can be measured simultaneously with propidium iodide (PI) staining of nuclei as an index of hypoxia-induced membrane damage. Hypoxia in rat proximal tubules is associated with a significant rise in [Caz+Ii, which precedes evidence of membrane damage as assessed by PI staining. This rise in [Caz+Ii activates calpain, a Ca2+-dependent cysteine protease and constitutive nitric oxide synthase (NOS), the CaZ+ dependent form of NOS. Inhibition of calpain with mechanistically and chemically dissimilar inhibitors provides marked cytoprotection against hypoxic and ionomycin induced proximal tubular injury. Similarly, inhibition of NOS with L-NAME, acidosis, tetrahydrobiopterin depletion and low extracellular calcium is associated with marked cytoprotection against hypoxic cellular injury. These observations are consistent with the early rise in [Ca"], initiating hypoxic injury by activating NOS and calpain.