Michael O’Connor

Overexpression of Hsp20 prevents endotoxin-induced myocardial dysfunction and apoptosis via inhibition of NF-κB activation

The occurrence of cardiovascular dysfunction in sepsis is associated with a significantly increased mortality rate of 70% to 90% compared with 20% in septic patients without cardiovascular impairment. Thus, rectification or blockade of myocardial depressant factors should partly ameliorate sepsis progression. Heat shock protein 20 (Hsp20) has been shown to enhance myocardial contractile function and protect against doxorubicin-induced cardiotoxicity. To investigate the possible role of Hsp20 in sepsis-mediated cardiac injury, we first examined the expression profiles of five major Hsps in response to lipopolysaccharide (LPS) challenge, and observed that only the expression of Hsp20 was downregulated in LPS-treated myocardium, suggesting that this decrease might be one of the mechanisms contributing to LPS-induced cardiovascular defects. Further studies using loss-of-function and gain-of-function approaches in adult rat cardiomyocytes verified that reduced Hsp20 levels were indeed correlated with the impaired contractile function. In fact, overexpression of Hsp20 significantly enhanced cardiomyocyte contractility upon LPS treatment. Moreover, after administration of LPS (25 microg/g) in vivo, Hsp20 transgenic mice (10-fold overexpression) displayed: 1) an improvement in myocardial function; 2) reduced the degree of cardiac apoptosis; and 3) decreased NF-kappaB activity, accompanied with reduced myocardial cytokines IL-1beta and TNF-alpha production, compared to the LPS-treated non-transgenic littermate controls. Thus, the increases in Hsp20 levels can protect against LPS-induced cardiac apoptosis and dysfunction, associated with inhibition of NF-kappaB activity, suggesting that Hsp20 may be a new therapeutic agent for the treatment of sepsis.

Lung injury after hemorrhage is age-dependent: role of peroxisome proliferator activated receptor γ

Objective:The incidence of multiple organ failure in pediatric trauma victims is lower than in the adult population. However, the molecular mechanisms are not yet defined. We investigated whether the pathophysiologic characteristics of hemorrhage-induced lung injury may be age dependent and may be regulated by the peroxisome proliferator-activated receptor &ggr; (PPAR&ggr;). Design:Prospective, laboratory investigation that used an established rodent model of hemorrhagic shock. Setting:University hospital laboratory. Subjects:Young (n = 67; 3–5 months old) and mature (n = 66; 11–13 months old) male rats. Interventions:Hemorrhagic shock was induced in young and mature rats by withdrawing blood to a mean arterial blood pressure of 50 mm Hg. After 3 hours, rats were rapidly resuscitated by infusing the shed blood and killed 3 hours thereafter. Measurements and Main Results:In young rats, lung injury was characterized by accumulation of red cells and neutrophils at the end of the resuscitation period; on Western blot analysis, lung expression of intercellular adhesion molecule-1 was increased. In contrast, the severity of lung injury was more pronounced in mature rats. Lung myeloperoxidase activity and expression of constitutive and inducible intercellular adhesion molecule-1 was significantly higher in mature rats compared with young rats. Mature rats also had higher plasma levels of cytokines and chemokines compared with young rats. This heightened inflammation was associated with higher degree of activation of nuclear factor-&kgr;B and down-regulation of PPAR&ggr; and heat shock factor-1 in the lung of mature rats compared with young rats. Treatment with the PPAR&ggr; ligand, the cyclopentenone prostaglandin 15-deoxy-&Dgr;12,14-prostaglandin J2, ameliorated lung injury in young, but not in mature animals. Conclusions:Lung injury after severe hemorrhage is age dependent and may be secondary to a diverse regulation of PPAR&ggr;.

The AMPK Activator Aicar Ameliorates Age-Dependent Myocardial Injury in Murine Hemorrhagic Shock.

ABSTRACT The development of myocardial dysfunction in patients with hemorrhagic shock is significantly impacted by the patient age. AMP-activated protein kinase (AMPK) is a pivotal orchestrator of energy homeostasis, which coordinates metabolic recovery after cellular stress. We investigated whether AMPK-regulated pathways are age-dependent in hemorrhage-induced myocardial injury and whether AMPK activation by 5-amino-4-imidazolecarboxamide riboside (AICAR) affords cardioprotective effects. Anesthetized C57/BL6 young (3–5 months old) and mature (9–12 months old) male mice were subjected to hemorrhagic shock by blood withdrawing followed by resuscitation with shed blood and Lactated Ringers solution. Mice were sacrificed at 3 h after resuscitation, and plasma and hearts were harvested for biochemical assays. Vehicle-treated mature mice exhibited higher myocardial injury and higher levels of plasma biomarkers of cardiovascular injury (endocan and follistatin) when compared with young mice. Cardiac cell mitochondrial structure was also markedly impaired in vehicle-treated mature mice when compared with young mice. At molecular analysis, an increase of the phosphorylated catalytic subunit pAMPK&agr; was associated with nuclear translocation of the peroxisome proliferator-activated receptor &ggr; coactivator-&agr; in young, but not mature mice. No changes in autophagy were observed as evaluated by the conversion of the light-chain (LC)3B-I protein to LC3B-II form. Treatment with AICAR ameliorated myocardial damage in both age groups. However, AICAR therapeutic effects were less effective in mature mice than young mice and involved distinct mechanisms of action. Thus, our data demonstrate that during hemorrhagic shock AMPK-dependent metabolic mechanisms are important for mitigating myocardial injury. However, these mechanisms are less competent with age.

Combined zinc supplementation with proinsulin C-peptide treatment decreases the inflammatory response and mortality in murine polymicrobial sepsis.

ABSTRACT Zinc is a trace element vital for immune function during host response to infection. The proinsulin C-peptide has been shown to exert beneficial effects through activation of the anti-inflammatory peroxisome proliferator–activated receptor &ggr; (PPAR&ggr;) in experimental endotoxemia. Some in vitro activities of C-peptide appear dependent on the presence of zinc. We investigated the effect of zinc supplementation before onset of sepsis on the anti-inflammatory properties of C-peptide. Male C57BL/6 mice were subjected to polymicrobial sepsis by cecal ligation and puncture (CLP). Mice received zinc gluconate (1.3 mg/kg) intraperitoneally (i.p.) for 3 days before CLP. One hour after CLP, animals received C-peptide (280 nmol/kg i.p.) or the antimicrobial agent imipenem (25 mg/kg i.p.). Cecal ligation and puncture was associated with an 11% survival rate, pulmonary leukosequestration, and liver injury. Molecular analysis in lungs of septic mice showed increased nuclear activation of the proinflammatory extracellular signal–regulated kinases 1 and 2 and nuclear factor &kgr;B, but decreased PPAR&ggr; expression, when compared with sham animals. Combination of zinc supplementation with C-peptide posttreatment significantly improved survival rate (61%) similarly to antibiotic treatment (60%), ameliorated lung architecture and liver function, reduced tissue neutrophil infiltration, and increased bacterial clearance when compared with vehicle, C-peptide, or zinc treatment alone. These beneficial effects were associated with restored lung nuclear expression of PPAR&ggr; and reduction of phosphorylated extracellular signal–regulated kinases 1 and 2 and nuclear factor &kgr;B activities in comparison to vehicle or single treatment protocols. Our data demonstrate that short-term zinc prophylaxis before the infectious insult is a requisite for the anti-inflammatory properties of C-peptide by facilitating modulation of inflammatory pathways.

Age-Dependent Changes in AMPK Metabolic Pathways in the Lung in a Mouse Model of Hemorrhagic Shock

&NA; The development of multiple organ failure in patients with hemorrhagic shock is significantly influenced by patient age. Adenosine monophosphate‐activated protein kinase (AMPK) is a crucial regulator of energy homeostasis, which coordinates metabolic repair during cellular stress. We investigated whether AMPK‐regulated signaling pathways are age‐dependent in hemorrhage‐induced lung injury and whether AMPK activation by 5‐amino‐4‐imidazole carboxamide riboside (AICAR) affords lung protective effects. Male C57/BL6 young mice (3‐5 mo), mature adult mice (9‐12 mo), and young AMPK&agr;1 knockout mice (3‐5 mo) were subjected to hemorrhagic shock by blood withdrawing, followed by resuscitation with shed blood and lactated Ringers solution. Plasma proinflammatory cytokines were similarly elevated in C57/BL6 young and mature adult mice after hemorrhagic shock. However, mature adult mice exhibited more severe lung edema and neutrophil infiltration, and higher mitochondrial damage in alveolar epithelial type II cells, than did young mice. No change in autophagy was observed. At molecular analysis, the phosphorylation of the catalytic subunit AMPK&agr;1 was associated with nuclear translocation of peroxisome proliferator‐activated receptor &ggr; co‐activator‐&agr; in young, but not mature, adult mice. Treatment with AICAR ameliorated the disruption of lung architecture in mice of both ages; however, effects in mature adult mice were different than young mice and also involved inhibition of nuclear factor‐&kgr;B. In young AMPK&agr;1 knockout mice, AICAR failed to improve hypotension and lung neutrophil infiltration. Our data demonstrate that during hemorrhagic shock, AMPK‐dependent metabolic repair mechanisms are important for mitigating lung injury. However, these mechanisms are less competent with age.

Age-dependent therapeutic effects of liver X receptor-α activation in murine polymicrobial sepsis.

The severity of sepsis is significantly affected by advanced age; however, age-dependent molecular mechanisms of this susceptibility are unknown. Nuclear liver X receptor-α (LXRα) is a regulator of lipid metabolism with associated anti-inflammatory properties. Here, we investigated the role of LXRα in age-dependent lung injury and outcome of sepsis. Male C57BL/6, LXRα-deficient (LXRα−/−) and wild type (WT) (LXRα+/+) mice of different ages were subjected to sepsis by cecal ligation and puncture (CLP). In pharmacological studies, treatment with the LXRα ligand T0901317 reduced lung neutrophil infiltration in C57BL/6 mice aged from 1 to 8 mo when compared with vehicle-treated animals subjected to CLP. The LXRα ligand improved survival in young mice (2–3 mo old) but did not affect survival or neutrophil infiltration in mature adult mice (11–13 mo old). Immunoblotting revealed an age-dependent decrease of lung LXRα levels. Young LXRα−/− mice (2–3 mo old) exhibited earlier mortality than age-matched WT mice after CLP. Lung damage and neutrophil infiltration, lung activation of the pro-inflammatory NF-κB and plasma IL-6 levels were higher in LXRα−/− mice 18 h after CLP compared with LXRα+/+ mice. This study suggests that the anti-inflammatory properties of LXRα in sepsis are age-dependent and severely compromised in mature adult animals.

Severe Acute Hyponatremia as an Initial Presentation of Acute Intermittent Porphyria Triggered by a Subdermal Etonogestrel Implant

Acute Intermittent Porphyria (AIP) is one of several disorders that arise from enzymatic derangements in the biosynthetic pathway of the heme molecule. AIP is the most common of the acute porphyrias worldwide, with an estimated prevalence of approximately 5 per 100,000 people [1]. Specifically, AIP results from a defective copy in one of the two genes for Porphobilinogen (PBG) deaminase, leading to the inadequate synthesis of the normal enzyme. Typically, patients will be symptom-free until a precipitating factor causes increased transit through the porphyrin metabolic pathway, resulting in accumulation in Aminolevulinic Acid (ALA) and Porphobilinogen (PBG) (Figure 1) [2-4]. AIP is the second most common porphyria but can have variable expression making diagnosis difficult [5]. We describe a case of acute intermittent porphyria that presented with non-specific lower back pain and progressive acute hyponatremia complicated by seizure, acute encephalopathy, tachycardia, and hypertension.

1321: RELATIONSHIP OF CRRT CIRCUIT FAILURE WITH BLOOD PRODUCT TRANSFUSION

Crit Care Med 2016 • Volume 44 • Number 12 (Suppl.) kidney injury (AKI). The combined use of functional and structural biomarkers is a better diagnostic and prognostic factor for AKI. Methods: Prospective, observational, comparative and longitudinal cohort study conducted in the American British Cowdray Medical Center (ABC) ICU in the time period comprehended between March 2016 and March 2018. Structural and functional biomarkers were measured in patients above the age of 18 years old admitted consecutively into the ICU. A statistical analysis was done to evaluate the diagnostic and prognostic capacity obtained when combined markers of structural and functional damage were used. The trial was approved by the relevant ethics committee at the participating site. Results: 19 patients were analyzed; baseline characteristics were not significantly different between groups. A total of 10 patients (53%) showed renal damage. Of the 10 patients with AKI, 2 (20%) had persistent and serious AKI. Of the measured biomarkers during admission of patients with and without AKI, only NGAL and adjusted NGAL showed significant difference [48.3 (1.1-3103) vs 6 (1.6-19.2) p=0.09 for NGAL and 147 (22-9374) vs 41 (11-146) p=0.04 for NGAL/Cr, respectively]. When comparing patients with persistent vs non persistent AKI, the biomarkers that showed significant differences were serum cystatin c [1.08 (0.78-1.22) vs 0.64 (0.40-1.12) p=0.02] and microalbuminuria [171 (55997) vs 39 (7-79) p=0.01, respectively]. The patients with the highest number of functional and structural markers had more persistence and a more severe condition of the AKI. Conclusions: Of the biomarkers measured for diagnosing AKI only adjusted NGAL showed clinical significance. The combined use of serum cystatin c, adjusted NGAL and microalbuminuria as functional and structural biomarkers can predict persistent and severe AKI in patients with previous cardiac surgery.

1052: GENETIC DEFICIENCY OF AMPKΑ1 IMPAIRS ENERGY PRODUCTION AND EXACERBATES ORGAN INJURY IN SEPTIC MICE

Crit Care Med 2015 • Volume 43 • Number 12 (Suppl.) 30.1 ml/kg vs. OB 17 ml/kg, p<0.01) was lower in the OB group. In both groups, the drip rates were significantly lower in patients who failed versus those who did not. Conclusions: This study questions the efficacy of non-weight vasopressor dosing. Adequately dosing obese patients may result in faster obtainment of goal MAP and decreased use of secondary agents. Randomized trials are needed to validate these results and examine adverse event rates.