Activation of AMP-activated protein kinase during sepsis/inflammation improves survival by preserving cellular metabolic fitness.

The purpose was to determine the role of AMPK activation in the renal metabolic response to sepsis, the development of sepsis-induced acute kidney injury (AKI) and on survival. In a prospective experimental study, 167 10- to 12-week-old C57BL/6 mice underwent cecal ligation and puncture (CLP) and human proximal tubule epithelial cells (TEC; HK2) were exposed to inflammatory mix (IM), a combination of lipopolysaccharide (LPS) and high mobility group box 1 (HMGB1). Renal/TEC metabolic fitness was assessed by monitoring the expression of drivers of oxidative phosphorylation (OXPHOS), the rates of utilization of OXPHOS/glycolysis in response to metabolic stress, and mitochondrial function by measuring O2 consumption rates (OCR) and the membrane potential (Δψm ). Sepsis/IM resulted in AKI, increased mortality, and in renal AMPK activation 6-24 hours after CLP/IM. Pharmacologic activation of AMPK with 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) or metformin during sepsis improved the survival, while AMPK inhibition with Compound C increased mortality, impaired mitochondrial respiration, decreased OCR, and disrupted TEC metabolic fitness. AMPK-driven protection was associated with increased Sirt 3 expression and restoration of metabolic fitness. Renal AMPK activation in response to sepsis/IM is an adaptive mechanism that protects TEC, organs, and the host by preserving mitochondrial function and metabolic fitness likely through Sirt3 signaling.

Toll-like Receptor 4 Signaling on Dendritic Cells Suppresses Polymorphonuclear Leukocyte CXCR2 Expression and Trafficking via Interleukin 10 During Intra-abdominal Sepsis.

BACKGROUND: Toll-like receptor 4 (TLR4) is a critical receptor involved in the sensing of gram-negative bacterial infection. However, the roles of TLR4 in sepsis are cell-type specific. Dendritic cells (DCs) are known to play a central role in microbial detection, alerting the immune system to the presence of infection and coordinating adaptive immune response. The goal of this study was to investigate the impact of DC-specific TLR4 signaling on host defense against intra-abdominal polymicrobial sepsis. METHODS: C57BL/6, global Tlr4 knockout, cell-specific knockout control, and CD11c-specific Tlr4(-/-) mice underwent cecal ligation and puncture (CLP). RESULTS: Specific deletion of TLR4 on DCs in mice improved survival and enhanced bacterial clearance. Deletion of TLR4 on DCs was associated with lower levels of circulating interleukin 10 (IL-10), higher polymorphonuclear leukocyte (PMN) accumulation in the peritoneal cavity, and higher expression of chemokine (C-X-C motif) receptor 2 (CXCR2) on PMNs after CLP. In vitro studies of DC and neutrophil cocultures confirmed that TLR4-dependent secretion of IL-10 from DCs regulated neutrophil CXCR2 expression. CONCLUSIONS: Our data shed light on a previously unrecognized role for TLR4 signaling on DCs in driving IL-10 secretion during sepsis and, through this pathway, regulates PMN recruitment via suppression of CXCR2 expression.

The Microcirculatory Response to Endotoxemia and Resuscitation Is a Marker of Regional Renal Perfusion, Renal Metabolic Stress, and Tubular Injury.

Aims: We sought to investigate the relationship between macrohemodynamic resuscitation and microcirculatory parameters with the response of microcirculatory flow, tissue-specific parameters of metabolic stress and injury. We hypothesized that early resuscitation based on macrohemodynamic parameters does not prevent the development of organ dysfunction in a porcine model of endotoxemic shock, and that sublingual microcirculatory parameters are associated with markers of tissue metabolic stress and injury. Results: Both resuscitation groups had significant increases in creatinine and neutrophil gelatinase-associated lipocalin as compared with baseline. Neither the macrovascular response to endotoxemia or resuscitation, nor group allocation predicted the development of acute kidney injury (AKI). Only a microvascular flow index (MFI) <2.5 was associated with the development of renal tubular injury and AKI, and with increased renal, liver, peritoneal, and sublingual lactate/pyruvate (L/P) ratio and lactate. Among systemic parameters, only partial pressure of carbon dioxide (PCO2) gap >6 and P(a-v)CO2/C(v-a)O2 >1.8 were associated with increased organ L/P ratio and AKI. Innovation and Conclusion: Our findings demonstrate that targeting macrohemodynamics to guide resuscitation during endotoxemic shock failed to predict tissue metabolic stress and the response of the microvasculature to resuscitation, and was unsuccessful in preventing tubular injury and AKI. Mechanistically, our data suggest that loss of hemodynamic coherence and decoupling of microvascular flow from tissue metabolic demand during endotoxemia may explain the lack of association between macrohemodynamics and perfusion goals. Finally, we demonstrate that MFI, PCO2 gap, and P(v-a)CO2/C(a-v)O2 ratio outperformed macrohemodynamic parameters at predicting the development of renal metabolic stress and tubular injury, and therefore, that these indices merit further validation as promising resuscitation targets. Antioxid. Redox Signal. 35, 1407-1425.

Tissue Inhibitor of Metalloproteinases-2 Mediates Kidney Injury during Sepsis.

INTRODUCTION: Urinary tissue inhibitor of metalloproteinases (TIMP)-2 has been identified as a predictive marker for acute kidney injury (AKI), including sepsis-associated AKI (S-AKI). Whether TIMP-2 might be causally related to AKI and hence represent a viable drug target is unclear. OBJECTIVE: The aim of this study was to evaluate whether suppression of TIMP-2 attenuates S-AKI. METHODS: Balb/c mice were randomized to sham or cecal ligation and puncture surgery and treated with or without a TIMP-2-neutralizing antibody. Animals were followed for 48 h and then sacrificed for analysis of TIMP-2 expression, cell cycle, and histology. RESULTS: Anti-TIMP-2 resulted in decreased lumen TIMP-2 expression which markedly increased cell cycle progression and attenuated epithelial cell injury by histology. CONCLUSIONS: TIMP-2 mediates S-AKI and appears to be a viable drug target.

Time-dependent effects of histone deacetylase inhibition in sepsis-associated acute kidney injury.

BACKGROUND: Sepsis, a dysregulated host response to infection with results in organ dysfunction, has been a major challenge to the development of effective therapeutics. Sepsis-associated acute kidney injury (S-AKI) results in a 3-5-fold increase in the risk of hospital mortality compared to sepsis alone. The development of therapies to reverse S-AKI could therefore significantly affect sepsis outcomes. However, the translation of therapies from preclinical studies into humans requires model systems that recapitulate clinical scenarios and the development of renal fibrosis indicative of the transition from acute to chronic kidney disease. RESULTS: Here we characterized a murine model of S-AKI induced by abdominal sepsis developing into a chronic phenotype. We applied a small molecule histone deacetylase-8 inhibitor, UPHD186, and found that early treatment, beginning at 48 h post-sepsis, worsened renal outcome accompanied by decreasing mononuclear cell infiltration in the kidney, skewing cells into a pro-inflammatory phenotype, and increased pro-fibrotic gene expression, while delayed treatment, beginning at 96 h post-sepsis, after the acute inflammation in the kidney had subsided, resulted in improved survival and kidney histology presumably through promoting proliferation and inhibiting fibrosis. CONCLUSIONS: These findings not only present a clinically relevant S-AKI model, but also introduce a timing dimension into S-AKI therapeutic interventions that delayed treatment with UPHD186 may enhance renal histologic repair. Our results provide novel insights into successful repair of kidney injury and sepsis therapy.

Fixed volume or fixed pressure: a murine model of hemorrhagic shock.

It is common knowledge that severe blood loss and traumatic injury can lead to a cascade of detrimental signaling events often resulting in mortality. These signaling events can also lead to sepsis and/or multiple organ dysfunction (MOD). It is critical then to investigate the causes of suppressed immune function and detrimental signaling cascades in order to develop more effective ways to help patients who suffer from traumatic injuries. This fixed pressure Hemorrhagic Shock (HS) procedure, although technically challenging, is an excellent resource for investigation of these pathophysiologic conditions. Advances in the assessment of biological systems, i.e. Systems Biology have enabled the scientific community to further understand complex physiologic networks and cellular communication patterns. (14) Hemorrhagic Shock has proven to be a vital tool for unveiling these cellular communication patterns as they relate to immune function. This procedure can be mastered! This procedure can also be used as either a fixed volume or fixed pressure approach. We adapted this technique in the murine model to enhance research in innate and adaptive immune function. Due to their small size HS in mice presents unique challenges. However due to the many available mouse strains, this species represents an unparalleled resource for the study of the biologic responses. The HS model is an important model for studying cellular communication patterns and the responses of systems such as hormonal and inflammatory mediator systems, and danger signals, i.e. DAMP and PAMP upregulation as it elicits distinct responses that differ from other forms of shock. The development of transgenic murine strains and the induction of biologic agents to inhibit specific signaling have presented valuable opportunities to further elucidate our understanding of the up and down regulation of signal transduction after severe blood loss, i.e. HS and trauma. There are numerous resuscitation methods (R) in association with HS and trauma. A fixed volume resuscitation method of solely lactated ringer solution (LR), equal to three times the shed blood volume, is used in this model to study endogenous mechanisms such as remote organ injury and systemic inflammation. This method of resuscitation is proven to be effective in evaluating the effects of HS and trauma.