The role of adenosine 1a receptor signaling on GFR early after the induction of sepsis

Sepsis and acute kidney injury (AKI) synergistically increase morbidity and mortality in the ICU. How sepsis reduces glomerular filtration rate (GFR) and causes AKI is poorly understood; one proposed mechanism includes tubuloglomerular feedback (TGF). When sodium reabsorption by the proximal tubules is reduced in normal animals, the macula densa senses increased luminal sodium chloride, and then adenosine-1a receptor (A1aR) signaling triggers tubuloglomerular feedback, reducing GFR through afferent arteriole vasoconstriction. We measured GFR and systemic hemodynamics early during cecal ligation and puncture-induced sepsis in wild-type and A1aR-knockout mice. A miniaturized fluorometer was attached to the back of each mouse and recorded the clearance of FITC-sinistrin via transcutaneous fluorescence to monitor GFR. Clinical organ injury markers and cytokines were measured and hemodynamics monitored using implantable transducer telemetry devices. In wild-type mice, GFR was stable within 1 h after surgery, declined by 43% in the next hour, and then fell to less than 10% of baseline after 2 h and 45 min. In contrast, in A1aR-knockout mice GFR was 37% below baseline immediately after surgery and then gradually declined over 4 h. A1aR-knockout mice had similar organ injury and inflammatory responses, albeit with lower heart rate. We conclude that transcutaneous fluorescence can accurately monitor GFR and detect changes rapidly during sepsis. Tubuloglomerular feedback plays a complex role in sepsis; initially, TGF helps maintain GFR in the 1st hour, and over the subsequent 3 h, TGF causes GFR to plummet. By 18 h, TGF has no cumulative effect on renal or extrarenal organ damage.

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Toll-like Receptor 7 Contributes to Inflammation, Organ Injury, and Mortality in Murine Sepsis

Anesthesiology ◽

10.1097/aln.0000000000002706 ◽

2019 ◽

Vol 131 (1) ◽

pp. 105-118 ◽

Cited By ~ 3

Author(s):

Wenling Jian ◽

Lili Gu ◽

Brittney Williams ◽

Yan Feng ◽

Wei Chao ◽

...

Keyword(s):

Acute Kidney Injury ◽

Immune Responses ◽

Knockout Mice ◽

Cecal Ligation ◽

Kidney Injury ◽

Innate Immune ◽

Organ Injury ◽

Innate Immune Responses ◽

Toll Like Receptor ◽

Wild Type

Abstract Editor’s Perspective What We Already Know about This Topic What This Article Tells Us That Is New Background Sepsis remains a critical illness with high mortality. The authors have recently reported that mouse plasma RNA concentrations are markedly increased during sepsis and closely associated with its severity. Toll-like receptor 7, originally identified as the sensor for single-stranded RNA virus, also mediates host extracellular RNA-induced innate immune responses in vitro and in vivo. Here, the authors hypothesize that innate immune signaling via Toll-like receptor 7 contributes to inflammatory response, organ injury, and mortality during polymicrobial sepsis. Methods Sepsis was created by (1) cecal ligation and puncture or (2) stool slurry peritoneal injection. Wild-type and Toll-like receptor 7 knockout mice, both in C57BL/6J background, were used. The following endpoints were measured: mortality, acute kidney injury biomarkers, plasma and peritoneal cytokines, blood bacterial loading, peritoneal leukocyte counts, and neutrophil phagocytic function. Results The 11-day overall mortality was 81% in wild-type mice and 48% in Toll-like receptor 7 knockout mice after cecal ligation and puncture (N = 27 per group, P = 0.0031). Compared with wild-type septic mice, Toll-like receptor 7 knockout septic mice also had lower sepsis severity, attenuated plasma cytokine storm (wild-type vs. Toll-like receptor 7 knockout, interleukin-6: 43.2 [24.5, 162.7] vs. 4.4 [3.1, 12.0] ng/ml, P = 0.003) and peritoneal inflammation, alleviated acute kidney injury (wild-type vs. Toll-like receptor 7 knockout, neutrophil gelatinase-associated lipocalin: 307 ± 184 vs.139 ± 41-fold, P = 0.0364; kidney injury molecule-1: 40 [16, 49] vs.13 [4, 223]-fold, P = 0.0704), lower bacterial loading, and enhanced leukocyte peritoneal recruitment and phagocytic activities at 24 h. Moreover, stool slurry from wild-type and Toll-like receptor 7 knockout mice resulted in similar level of sepsis severity, peritoneal cytokines, and leukocyte recruitment in wild-type animals after peritoneal injection. Conclusions Toll-like receptor 7 plays an important role in the pathogenesis of polymicrobial sepsis by mediating host innate immune responses and contributes to acute kidney injury and mortality.

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Nutritional Effects on Host Response to Lung Infections with Mucoid Pseudomonas aeruginosa in Mice

Infection and Immunity ◽

10.1128/iai.72.3.1479-1486.2004 ◽

2004 ◽

Vol 72 (3) ◽

pp. 1479-1486 ◽

Cited By ~ 15

Author(s):

Anna M. van Heeckeren ◽

Mark Schluchter ◽

Lintong Xue ◽

Juan Alvarez ◽

Steven Freedman ◽

...

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Docosahexaenoic Acid ◽

Knockout Mice ◽

Host Response ◽

Inflammatory Responses ◽

Liquid Diet ◽

Wild Type ◽

Agarose Beads ◽

Lung Infections

ABSTRACT In cystic fibrosis, a recessive genetic disease caused by defects in the cystic fibrosis conductance regulator (CFTR), the main cause of death is lung infection and inflammation. Nutritional deficits have been proposed to contribute to the excessive host inflammatory response in both humans and Cftr-knockout mice. Cftr-knockout mice and gut-corrected Cftr-knockout mice expressing human CFTR primarily in the gut were challenged with Pseudomonas aeruginosa-laden agarose beads; they responded similarly with respect to bronchoalveolar lavage cell counts and levels of the acute-phase cytokines tumor necrosis factor alpha, interleukin-1β (IL-1β), and IL-6. Wild-type mice fed the liquid diet used to prevent intestinal obstruction in Cftr-knockout mice had inflammatory responses to P. aeruginosa-laden agarose beads similar to those of wild-type mice fed an enriched solid diet, so dietary effects are unlikely to account for differences between wild-type mice and mice with cystic fibrosis. Finally, since cystic fibrosis patients and Cftr-knockout mice have an imbalance in fatty acids (significantly lower-than-normal levels of docosahexaenoic acid), the effects of specific supplementation with docosahexaenoic acid of wild-type and Cftr-knockout mice on their inflammatory responses to P. aeruginosa-laden agarose beads were tested. There were no significant differences (P = 0.35) in cumulative survival rates between Cftr-knockout mice and wild-type mice provided with either the liquid diet Peptamen or Peptamen containing docosahexaenoic acid. In conclusion, diet and docosahexaenoic acid imbalances alone are unlikely to explain the differences in the host response to lung infections with mucoid P. aeruginosa between mice with cystic fibrosis and their wild-type counterparts.

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Comparison of serum creatinine and serum cystatin C as biomarkers to detect sepsis-induced acute kidney injury and to predict mortality in CD-1 mice

AJP Renal Physiology ◽

10.1152/ajprenal.00025.2013 ◽

2014 ◽

Vol 307 (8) ◽

pp. F939-F948 ◽

Cited By ~ 26

Author(s):

Asada Leelahavanichkul ◽

Ana Carolina P. Souza ◽

Jonathan M. Street ◽

Victor Hsu ◽

Takayuki Tsuji ◽

...

Keyword(s):

Acute Kidney Injury ◽

Serum Creatinine ◽

Cystatin C ◽

Cecal Ligation ◽

Kidney Injury ◽

Organ Damage ◽

Experimental Sepsis ◽

Sepsis Mortality ◽

Serum Cystatin C ◽

Serum Cystatin

Acute kidney injury (AKI) dramatically increases sepsis mortality, but AKI diagnosis is delayed when based on serum creatinine (SCr) changes, due in part, to decreased creatinine production. During experimental sepsis, we compared serum cystatin C (sCysC), SCr, and blood urea nitrogen (BUN) to inulin glomerular filtration rate (iGFR) before or 3–18 h after cecal ligation and puncture (CLP)-induced sepsis in CD-1 mice. sCysC had a faster increase and reached peak levels more rapidly than SCr in both sepsis and bilateral nephrectomy (BiNx) models. sCysC was a better surrogate of iGFR than SCr during sepsis. Combining sCysC with SCr values into a composite biomarker improved correlation with iGFR better than any biomarker alone or any other combination. We determined the renal contribution to sCysC handling with BiNx. sCysC and SCr were lower post-BiNx/CLP than post-BiNx alone, despite increased inflammatory and nonrenal organ damage biomarkers. Sepsis decreased CysC production in nephrectomized mice without changing body weight or CysC space. Sepsis decreased sCysC production and increased nonrenal clearance, similar to effects of sepsis on SCr. sCysC, SCr, and BUN were measured 6 h postsepsis to link AKI with mortality. Mice with above-median sCysC, BUN, or SCr values 6 h postsepsis died earlier than mice with below-median values, corresponding to a substantial AKI association with sepsis mortality in this model. sCysC performs similarly to SCr in classifying mice at risk for early mortality. We conclude that sCysC detects AKI early and better reflects iGFR in CLP-induced sepsis. This study shows that renal biomarkers need to be evaluated in specific contexts.

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Disruption of CXCR6 Ameliorates Kidney Inflammation and Fibrosis in Deoxycorticosterone Acetate/Salt Hypertension

Scientific Reports ◽

10.1038/s41598-019-56933-7 ◽

2020 ◽

Vol 10 (1) ◽

Cited By ~ 2

Author(s):

Yuanbo Wu ◽

Changlong An ◽

Xiaogao Jin ◽

Zhaoyong Hu ◽

Yanlin Wang

Keyword(s):

Knockout Mice ◽

Critical Role ◽

Kidney Injury ◽

Wild Type ◽

Salt Treatment ◽

Hypertensive Nephropathy ◽

Deoxycorticosterone Acetate ◽

Kidney Fibrosis ◽

Salt Hypertension

AbstractCirculating cells have a pathogenic role in the development of hypertensive nephropathy. However, how these cells infiltrate into the kidney are not fully elucidated. In this study, we investigated the role of CXCR6 in deoxycorticosterone acetate (DOCA)/salt-induced inflammation and fibrosis of the kidney. Following uninephrectomy, wild-type and CXCR6 knockout mice were treated with DOCA/salt for 3 weeks. Blood pressure was similar between wild-type and CXCR6 knockout mice at baseline and after treatment with DOCA/salt. Wild-type mice develop significant kidney injury, proteinuria, and kidney fibrosis after three weeks of DOCA/salt treatment. CXCR6 deficiency ameliorated kidney injury, proteinuria, and kidney fibrosis following treatment with DOCA/salt. Moreover, CXCR6 deficiency inhibited accumulation of bone marrow–derived fibroblasts and myofibroblasts in the kidney following treatment with DOCA/salt. Furthermore, CXCR6 deficiency markedly reduced the number of macrophages and T cells in the kidney after DOCA/salt treatment. In summary, our results identify a critical role of CXCR6 in the development of inflammation and fibrosis of the kidney in salt-sensitive hypertension.

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A1 adenosine receptor knockout mice exhibit increased mortality, renal dysfunction, and hepatic injury in murine septic peritonitis

AJP Renal Physiology ◽

10.1152/ajprenal.00470.2004 ◽

2005 ◽

Vol 289 (2) ◽

pp. F369-F376 ◽

Cited By ~ 57

Author(s):

George Gallos ◽

Thomas D. Ruyle ◽

Charles W. Emala ◽

H. Thomas Lee

Keyword(s):

Organ Dysfunction ◽

Knockout Mice ◽

Adenosine Receptor ◽

Ischemia Reperfusion ◽

Cecal Ligation ◽

Intercellular Adhesion Molecule ◽

Myeloperoxidase Activity ◽

Wild Type ◽

Wild Type Littermate ◽

Septic Peritonitis

Sepsis is a leading cause of multiorgan dysfunction and death in hospitalized patients. Dysregulated inflammatory processes and apoptosis contribute to the pathogenesis of sepsis-induced organ dysfunction and death. A1 adenosine receptor (A1AR) activation reduces inflammation and apoptosis after ischemia-reperfusion injury. Therefore, we questioned whether A1AR-mediated reduction of inflammation and apoptosis could improve mortality and organ dysfunction in a murine model of sepsis. A1AR knockout mice (A1 knockout) and their wild-type (A1 wild-type) littermate controls were subjected to cecal ligation and double puncture (CLP) with a 20-gauge needle. A1 knockout mice or A1 wild-type mice treated with 1,3-dipropyl-8-cyclopentylxanthine (a selective A1AR antagonist) had a significantly higher mortality rate compared with A1 wild-type mice following CLP. Mice lacking endogenous A1ARs demonstrated significant elevations in plasma creatinine, alanine aminotransferase, aspartate aminotransferase, keratinocyte-derived chemokine, and tumor necrosis factor-α 24 h after induction of sepsis compared with wild-type mice. The renal corticomedullary junction from A1 knockout mice also exhibited increased myeloperoxidase activity, intercellular adhesion molecule-1 protein, and mRNA encoding proinflammatory cytokines compared with renal samples from A1 wild-type littermate controls. No difference in renal tubular apoptosis was detected between A1 knockout and A1 wild-type mice. We conclude that endogenous A1AR activation confers a protective effect in mice from septic peritonitis primarily by attenuating the hyperacute inflammatory response in sepsis.

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Differential effect of imipenem treatment on injury caused by cecal ligation and puncture in wild-type and NK cell-deficient β2-microgloblin knockout mice

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00338.2005 ◽

2006 ◽

Vol 290 (2) ◽

pp. G277-G284 ◽

Cited By ~ 6

Author(s):

Victor T. Enoh ◽

Cheng Y. Lin ◽

Tushar K. Varma ◽

Edward R. Sherwood

Keyword(s):

Metabolic Acidosis ◽

Knockout Mice ◽

Nk Cell ◽

Plasma Concentrations ◽

Cecal Ligation ◽

Systemic Infection ◽

Cecal Ligation And Puncture ◽

Wild Type ◽

Term Survival ◽

The Impact

Our previous studies showed that β2-microglobulin knockout mice treated with anti-asialoGM1 (β2MKO/αAsGM1 mice) are resistant to injury caused by cecal ligation and puncture (CLP). However, CLP-induced injury is complex. Potential mechanisms of injury include systemic infection, cecal ischemia, and translocation of bacterial toxins such as endotoxin and superantigens. Currently, it is unclear which of these mechanisms of injury contributes to mortality in wild-type mice and whether β2MKO/αAsGM1 mice are resistant to any particular mechanisms of injury. In the present study, we hypothesized that systemic infection is the major cause of injury after CLP in wild-type mice and that β2MKO/αAsGM1 mice are resistant to infection-induced injury. To test this hypothesis, wild-type and β2MKO/αAsGM1 mice were treated with the broad-spectrum antibiotic imipenem immediately after CLP to decrease the impact of systemic infection in our model. Treatment of wild-type and β2MKO/αAsGM1 mice with imipenem decreased bacterial counts by at least two orders of magnitude. However, all wild-type mice, whether treated with saline or imipenem, died by 42 h after CLP and had significant hypothermia, metabolic acidosis, and high plasma concentrations of the cytokines interleukin-6, macrophage inflammatory protein-2, and keratinocyte-derived chemokine. β2MKO/αAsGM1 mice showed 40% long-term survival, which was increased to 90% by imipenem treatment. β2MKO/αAsGM1 mice had less hypothermia, decreased metabolic acidosis, and lower cytokine concentrations at 18 h after CLP compared with wild-type mice. These results suggest that infection is not the major cause of mortality for wild-type mice in our model of CLP. Other mechanisms of injury such as cecal ischemia or translocation of microbial toxins may be more important. β2MKO/αAsGM1 mice appear resistant to these early, non-infection-related causes of CLP-induced injury but showed delayed mortality associated with bacterial dissemination, which was ablated by treatment with imipenem.

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Transient Receptor Potential Melastatin 2 Protects Mice against Polymicrobial Sepsis by Enhancing Bacterial Clearance

Anesthesiology ◽

10.1097/aln.0000000000000275 ◽

2014 ◽

Vol 121 (2) ◽

pp. 336-351 ◽

Cited By ~ 30

Author(s):

XiaoWei Qian ◽

Tomohiro Numata ◽

Kai Zhang ◽

CaiXia Li ◽

JinChao Hou ◽

...

Keyword(s):

Bone Marrow ◽

Knockout Mice ◽

Transient Receptor Potential ◽

Cecal Ligation ◽

Polymicrobial Sepsis ◽

Bacterial Clearance ◽

Cecal Ligation And Puncture ◽

Bacterial Burden ◽

Wild Type ◽

Transient Receptor Potential Melastatin

Abstract Background: Recent studies suggest that the transient receptor potential melastatin 2 (TRPM2) channel plays an important role in inflammation and immune response. However, the role and mechanism of TRPM2 in polymicrobial sepsis remain unclear. Methods: The authors explored the effects of genetic disruption of TRPM2 on mortality (n = 15), bacterial clearance (n = 6), organ injury, and systemic inflammation during cecal ligation and puncture–induced sepsis. Electrophysiology, immunoblot, bacterial clearance experiment, and quantitative real-time polymerase chain reaction were used to explore the role and mechanism of TRPM2 in sepsis. Results: After cecal ligation and puncture, Trpm2-knockout mice had increased mortality compared with wild-type mice (73.3 vs. 40%, P = 0.0289). The increased mortality was associated with increased bacterial burden, organ injury, and systemic inflammation. TRPM2-mediated Ca2+ influx plays an important role in lipopolysaccharide or cecal ligation and puncture–induced heme oxygenase-1 (HO-1) expression in macrophage. HO-1 up-regulation decreased bacterial burden both in wild-type bone marrow–derived macrophages and in cecal ligation and puncture–induced septic wild-type mice. Disruption of TRPM2 decreased HO-1 expression and increased bacterial burden in bone marrow–derived macrophages. Pretreatment of Trpm2-knockout bone marrow–derived macrophages with HO-1 inducer markedly increased HO-1 expression and decreased bacterial burden. Pretreatment of Trpm2-knockout mice with HO-1 inducer reversed the susceptibility of Trpm2-knockout mice to sepsis by enhancing the bacterial clearance. In addition, septic patients with lower monocytic TRPM2 and HO-1 messenger RNA levels had a worse outcome compared with septic patients with normal monocytic TRPM2 and HO-1 messenger RNA levels. TRPM2 levels correlated with HO-1 levels in septic patients (r = 0.675, P = 0.001). Conclusion: The study data demonstrate a protective role of TRPM2 in controlling bacterial clearance during polymicrobial sepsis possibly by regulating HO-1 expression.

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Toll-Like Receptors 2 and 4 Contribute to Sepsis-Induced Depletion of Spleen Dendritic Cells

Infection and Immunity ◽

10.1128/iai.00238-09 ◽

2009 ◽

Vol 77 (12) ◽

pp. 5651-5658 ◽

Cited By ~ 40

Author(s):

Frédéric Pène ◽

Emilie Courtine ◽

Fatah Ouaaz ◽

Benjamin Zuber ◽

Bertrand Sauneuf ◽

...

Keyword(s):

Dendritic Cells ◽

Knockout Mice ◽

Cecal Ligation ◽

Polymicrobial Sepsis ◽

Interleukin 12 ◽

Maturation Process ◽

Wild Type ◽

Cpg Dna ◽

Functional Features ◽

Secondary Lymphoid Organs

ABSTRACT Depletion of dendritic cells (DC) in secondary lymphoid organs is a hallmark of sepsis-induced immune dysfunction. In this setting, we investigated if Toll-like receptor (TLR)-dependent signaling might modulate the maturation process and the survival of DC. Using a model of sublethal polymicrobial sepsis induced by cecal ligation and puncture, we investigated the quantitative and functional features of spleen DC in wild-type, TLR2−/−, TLR4−/−, and TLR2−/− TLR4−/− mice. By 24 h, a decrease in the relative percentage of CD11chigh spleen DC occurred in wild-type mice but was prevented in TLR2−/−, TLR4−/−, and TLR2−/− TLR4−/− mice. In wild-type mice, sepsis dramatically affected both CD11c+ CD8α+ and CD11c+ CD8α− subsets. In all three types of knockout mice studied, the CD11c+ CD8α+ subset followed a depletion pattern similar to that for wild-type mice. In contrast, the loss of CD11c+ CD8α− cells was attenuated in TLR2−/− and TLR4−/− mice and completely prevented in TLR2−/− TLR4−/− mice. Accordingly, apoptosis of spleen DC was increased in septic wild-type mice and inhibited in knockout mice. In addition we characterized the functional features of spleen DC obtained from septic mice. As shown by increased expression of major histocompatibility complex class II and CD86, polymicrobial sepsis induced maturation of DC, with subsequent increased capacity to prime T lymphocytes, similarly in wild-type and knockout mice. In response to CpG DNA stimulation, production of interleukin-12 was equally impaired in DC obtained from wild-type and knockout septic mice. In conclusion, although dispensable for the DC maturation process, TLR2 and TLR4 are involved in the mechanisms leading to depletion of spleen DC following polymicrobial sepsis.

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Methyl-2-acetamidoacrylate, an ethyl pyruvate analog, decreases sepsis-induced acute kidney injury in mice

AJP Renal Physiology ◽

10.1152/ajprenal.90442.2008 ◽

2008 ◽

Vol 295 (6) ◽

pp. F1825-F1835 ◽

Cited By ~ 51

Author(s):

Asada Leelahavanichkul ◽

Hideo Yasuda ◽

Kent Doi ◽

Xuzhen Hu ◽

Hua Zhou ◽

...

Keyword(s):

Liver Injury ◽

Combination Therapy ◽

Ethyl Pyruvate ◽

Statistical Significance ◽

Cecal Ligation ◽

Kidney Injury ◽

Organ Damage ◽

Therapeutic Window ◽

Beneficial Effects ◽

Cytokine Levels

We tested the anti-inflammatory agent methyl-2-acetamidoacrylate (M2AA), an ethyl pyruvate analog, in a cecal ligation-and-puncture (CLP) model of sepsis in CD-1 mice. M2AA administration at the time of CLP improved survival, renal function, kidney histology, liver injury, and splenocyte apoptosis, and lowered cytokine levels (TNF-α, IL-6, IFN-γ, and IL-10). When M2AA treatment was delayed 6 h (but not 12 h), M2AA still significantly reduced kidney dysfunction, liver injury, splenocyte apoptosis, and cytokine levels. NF-κB, a M2AA target, was transiently activated in spleen, peaking at 6 h; kidney and liver NF-κB increased steadily with a plateau at 12–24 h. M2AA reduced NF-κB activation in spleen at 6 h and in kidney and liver at 24 h. Splenectomy diminished the ability of M2AA to reduce cytokines, especially IL-6, but M2AA still decreased kidney and liver dysfunction, suggesting that splenic NF-κB is not central to M2AA action. In contrast, beneficial effects of chloroquine on cytokines and organ damage were neutralized by splenectomy, demonstrating a spleen-specific chloroquine target. Because M2AA and chloroquine act differently, we tested this combination. Survival at 96 h was highest with combination therapy (57%) vs. chloroquine (38%), M2AA (47.6%), or vehicle (5%). The benefit of combination therapy over chloroquine or M2AA alone did not reach statistical significance, indicating potential mechanistic overlap. We conclude that the transient target(s) for M2AA responsible for the narrow 6-h therapeutic window is not splenic NF-κB. Identifying this new target and downstream signaling pathways could lengthen the therapeutic window and improve combination therapy with chloroquine.

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Villin and actin in the mouse kidney brush-border membrane bind to and are degraded by meprins, an interaction that contributes to injury in ischemia-reperfusion

AJP Renal Physiology ◽

10.1152/ajprenal.00703.2010 ◽

2011 ◽

Vol 301 (4) ◽

pp. F871-F882 ◽

Cited By ~ 21

Author(s):

Elimelda Moige Ongeri ◽

Odinaka Anyanwu ◽

W. Brian Reeves ◽

Judith S. Bond

Keyword(s):

Brush Border ◽

Knockout Mice ◽

Brush Border Membrane ◽

Ischemia Reperfusion ◽

Kidney Injury ◽

Mouse Kidney ◽

Cytoskeletal Proteins ◽

Kidney Tubules ◽

Wild Type

Meprins, metalloproteinases abundantly expressed in the brush-border membranes (BBMs) of rodent proximal kidney tubules, have been implicated in the pathology of renal injury induced by ischemia-reperfusion (IR). Disruption of the meprin β gene and actinonin, a meprin inhibitor, both decrease kidney injury resulting from IR. To date, the in vivo kidney substrates for meprins are unknown. The studies herein implicate villin and actin as meprin substrates. Villin and actin bind to the cytoplasmic tail of meprin β, and both meprin A and B are capable of degrading villin and actin present in kidney proteins as well as purified recombinant forms of these proteins. The products resulting from degradation of villin and actin were unique to each meprin isoform. The meprin B cleavage site in villin was Glu744-Val745. Recombinant forms of rat meprin B and homomeric mouse meprin A had Km values for villin and actin of ∼1 μM (0.6–1.2 μM). The kcat values varied substantially (0.6–128 s−1), resulting in different efficiencies for cleavage, with meprin B having the highest kcat/ Km values (128 M−1·s−1 × 106). Following IR, meprins and villin redistributed from the BBM to the cytosol. A 37-kDa actin fragment was detected in protein fractions from wild-type, but not in comparable preparations from meprin knockout mice. The levels of the 37-kDa actin fragment were significantly higher in kidneys subjected to IR. The data establish that meprins interact with and cleave villin and actin, and these cytoskeletal proteins are substrates for meprins.

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