Fong Lam

Hypertrophic cardiomyopathy and dysregulation of cardiac energetics in a mouse model of biliary fibrosis

Cardiac dysfunction is a major cause of morbidity and mortality in patients with end‐stage liver disease; yet the mechanisms remain largely unknown. We hypothesized that the complex interrelated impairments in cardiac structure and function secondary to progression of liver diseases involve alterations in signaling pathways engaged in cardiac energy metabolism and hypertrophy, augmented by direct effects of high circulating levels of bile acids. Biliary fibrosis was induced in male C57BL/6J mice by feeding a 0.1% 3,5‐diethoxycarbonyl‐1,4‐dihydroxychollidine (DDC) supplemented diet. After 3 weeks, mice underwent live imaging (dual energy x‐ray absorptiometry [DEXA] scanning, two‐dimensional echocardiography [2DE], electrocardiography, cardiac magnetic resonance imaging), exercise treadmill testing, and histological and biochemical analyses of livers and hearts. Compared with chow‐fed mice, DDC‐fed mice fatigued earlier on the treadmill, with reduced VO2. Marked changes were identified electrophysiologically (bradycardia and prolonged QT interval) and functionally (hyperdynamic left ventricular [LV] contractility along with increased LV thickness). Hearts of DDC‐fed mice showed hypertrophic signaling (activation of v‐akt murine thymoma viral oncogene/protein kinase B [AKT], inhibition of glycogen synthase kinase‐3β [GSK3β], a 20‐fold up‐regulation of β myosin heavy chain RNA and elevated Gsα/Giα ratio. Genes regulating cardiac fatty acid oxidation pathways were suppressed, along with a threefold increase in myocardial glycogen content. Treatment of mouse cardiomyocytes (which express the membrane bile acid receptor TGR5) with potent natural TGR5 agonists, taurochenodeoxycholic acid and lithocholic acid, activated AKT and inhibited GSK3β, similar to the changes seen in DDC‐fed mouse hearts. This provides support for a novel mechanism whereby circulating natural bile acids can induce signaling pathways in heart associated with hypertrophy. Conclusion: Three weeks of DDC feeding‐induced biliary fibrosis leads to multiple functional, metabolic, electrophysiological, and hypertrophic adaptations in the mouse heart, recapitulating some of the features of human cirrhotic cardiomyopathy. Hepatology 2010;51:2097–2107

Histone induced platelet aggregation is inhibited by normal albumin

INTRODUCTION Histones are small, nuclear proteins that serve to package DNA. Recent reports suggest that extracellular histones, including histone H4, may contribute to the pathogenesis of sepsis; they promote platelet aggregation and thrombosis when released into the circulation during inflammation or cell death. The mechanisms by which the body minimizes the deleterious effects of circulating histones are unclear. Because histones can bind to plasma proteins, including albumin, we hypothesized that normal albumin can prevent histones from activating platelets. MATERIALS AND METHODS Platelets and platelet-free plasma were obtained from healthy, adult subjects. The dose-dependent effects of histone H4 on platelet aggregation were studied by optical aggregometry. The effects of native and albumin-depleted plasma (prepared by affinity chromatography) on histone-induced platelet aggregation were also assessed. The effects of normal and surface-neutralized albumin (through modification of carboxyl groups) on histone-induced platelet activation and aggregation were evaluated using flow cytometry and aggregometry. RESULTS Histone H4 induced platelet aggregation in a dose-dependent manner. This histone-induced platelet aggregation was inhibited by both plasma and human serum albumin in a dose-dependent fashion. Furthermore, depletion of albumin from plasma reduced its ability to inhibit aggregation. Finally, surface neutralization of albumin decreased its ability to inhibit histone-induced activation and aggregation. DISCUSSION These data suggest that normal albumin serves a role in preventing histone-induced platelet aggregation in a charge-dependent manner.

Platelets and Their Interactions with Other Immune Cells.

Platelets are anucleate blood cells, long known to be critically involved in hemostasis and thrombosis. In addition to their role in blood clots, increasing evidence reveals significant roles for platelets in inflammation and immunity. However, the notion that platelets represent immune cells is not broadly recognized in the field of Physiology. This article reviews the role of platelets in inflammation and immune responses, and highlights their interactions with other immune cells, including examples of major functional consequences of these interactions.

Coinfection with Staphylococcus aureus increases risk of severe coagulopathy in critically ill children with influenza A (H1N1) virus infection.

Objectives:H1N1 influenza with coinfections has been implicated to have high morbidity and mortality. We hypothesized that critically ill children with 2009 H1N1 and coinfections are at a higher risk of developing disseminated intravascular coagulation. Design:The chart review included demographics, length-of-stay, severity of illness score (Pediatric Risk of Mortality III acute physiology score), clinical laboratories, and outcomes at hospital day 90 data. Patients were classified as having methicillin-sensitive or -resistant Staphylococcus aureus, other, or no coinfections. Setting:Single-center pediatric intensive care unit. Patients:Sixty-six consecutive patients with 2009 H1N1 and influenza A infection. Interventions:None. Main Results:There were 12, 22, and 32 patients with methicillin-sensitive or -resistant Staphylococcus aureus, other, and no coinfections, respectively. Pediatric critical care unit length-of-stay was 11, 10, and 5.5 days (median), and survival at day 90 was 83%, 96%, and 91% in patients with methicillin-sensitive or -resistant Staphylococcus aureus, other, and no coinfections. Patients with methicillin-sensitive or -resistant Staphylococcus aureus coinfections compared to patients with other, and no coinfections had higher Pediatric Risk of Mortality III acute physiology scores (14 [6–25] vs. 7 [2–10], p = .052 and 6 [2.5–10], p = .008; median [interquartile range]), higher D-dimer (16.1 [7.9–19.3] vs. 1.6 [1.1–4], p = .02 and 2.3 [0.8–8.7] µg/mL, p = .05), longer prothrombin time (19.3 [15.4–25.9] vs. 15.3 [14.8–17.1], p = .04 and 16.6 [14.7–20.4] secs, p < .39) at admission, and lower day-7 platelet counts (90K [26–161K] vs. 277K [98–314], p = .03 and 256K [152–339]/mm3, p < .07). Patients with methicillin-sensitive or -resistant Staphylococcus aureus coinfections compared to patients without coinfections were more likely to be sicker with Pediatric Risk of Mortality III acute physiology score >10 vs. <10 (relative risk 2.4; 95% confidence interval 1.2–4.7; p = .035) and have overt disseminated intravascular coagulation (relative risk 4.4; 95% confidence interval 1.3–15.8, p = .025). Conclusions:During the 2009–2010 H1N1 pandemic, pediatric patients with influenza A and methicillin-sensitive or -resistant Staphylococcus aureus coinfections were sicker and more likely to develop disseminated intravascular coagulation than patients with other or no coinfections.

Polymicrobial sepsis and endotoxemia promote microvascular thrombosis via distinct mechanisms

Summary.  Background: We reported recently that endotoxemia promotes microvascular thrombosis in cremaster venules of wild‐type mice, but not in mice deficient in toll‐like receptor 4 (TLR4) or von Willebrand factor (VWF). Objective: To determine whether the clinically relevant model of polymicrobial sepsis induced by cecal ligation/perforation (CLP) induces similar responses via the same mechanisms as endotoxemia. Methods: We used a light/dye‐injury model of thrombosis in the cremaster microcirculation of wild‐type mice and mice deficient in toll‐like receptor‐4 (C57BL/10ScNJ), toll‐like receptor 2 (TLR2), or VWF. Mice underwent CLP or sham surgery, or an intraperitoneal injection of endotoxin (LPS) or saline. In the CLP model, we assessed the influence of fluid replacement on thrombotic responses. Results: Both CLP and LPS enhanced thrombotic occlusion in wild‐type mice. In contrast to LPS, CLP enhanced thrombosis in TLR4‐ and VWF‐deficient strains. While TLR2‐deficient mice did not demonstrate enhanced thrombosis following CLP, LPS enhanced thrombosis in these mice. LPS, but not CLP, increased plasma VWF antigen relative to controls. Septic mice, particularly those undergoing CLP, developed significant hemoconcentration. Intravenous fluid replacement with isotonic saline prevented the hemoconcentration and prothrombotic responses to CLP, though fluids did not prevent the prothrombotic response to LPS. Conclusions: Polymicrobial sepsis induced by CLP and endotoxemia promote microvascular thrombosis via distinct mechanisms; enhanced thrombosis induced by CLP requires TLR2 but not TLR4 or VWF. The salutary effects of intravenous fluid replacement on microvascular thrombosis in polymicrobial sepsis remain to be characterized.

Histones stimulate von Willebrand factor release in vitro and in vivo.

Histones, nuclear proteins that normally package DNA in cells, are increasingly recognized as important mediators of inflammation and thrombosis, and have recently been linked to death in sepsis.1 They are released during inflammation by activated leukocytes, primarily neutrophils, as part of neutrophil extracellular traps (NETs).2 Although NETs play an important role in innate immunity, components within NETs, namely histones, are linked to platelet activation3 and the development of thrombi.4 Indeed, mice that are unable to produce NETs have decreased thrombus formation.5 Although most descriptions of the prothrombotic effects of histones are based on activation of platelets and fibrin formation, less is known about their effects on endothelial cells.

A homing system targets therapeutic T cells to brain cancer

Successful T cell immunotherapy for brain cancer requires that the T cells can access tumour tissues, but this has been difficult to achieve. Here we show that, in contrast to inflammatory brain diseases such as multiple sclerosis, where endothelial cells upregulate ICAM1 and VCAM1 to guide the extravasation of pro-inflammatory cells, cancer endothelium downregulates these molecules to evade immune recognition. By contrast, we found that cancer endothelium upregulates activated leukocyte cell adhesion molecule (ALCAM), which allowed us to overcome this immune-evasion mechanism by creating an ALCAM-restricted homing system (HS). We re-engineered the natural ligand of ALCAM, CD6, in a manner that triggers initial anchorage of T cells to ALCAM and conditionally mediates a secondary wave of adhesion by sensitizing T cells to low-level ICAM1 on the cancer endothelium, thereby creating the adhesion forces necessary to capture T cells from the bloodstream. Cytotoxic HS T cells robustly infiltrated brain cancers after intravenous injection and exhibited potent antitumour activity. We have therefore developed a molecule that targets the delivery of T cells to brain cancer.Therapeutic T cells bearing ligands engineered to optimize adhesion and transmigration through the blood–brain barrier can be targeted to brain tumours.

Recombinant Human Vimentin Binds to P-Selectin and Blocks Neutrophil Capture and Rolling on Platelets and Endothelium

Leukocyte adhesion to vascular endothelium and platelets is an early step in the acute inflammatory response. The initial process is mediated through P-selectin glycoprotein ligand-1 (PSGL-1) on leukocytes binding to platelets adhered to endothelium and the endothelium itself via P-selectin. Although these interactions are generally beneficial, pathologic inflammation may occur in undesirable circumstances, such as in acute lung injury (ALI) and ischemia and reperfusion injury. Therefore, the development of novel therapies to attenuate inflammation may be beneficial. In this article, we describe the potential benefit of using a recombinant human vimentin (rhVim) on reducing human leukocyte adhesion to vascular endothelium and platelets under shear stress. The addition of rhVim to whole blood and isolated neutrophils decreased leukocyte adhesion to endothelial and platelet monolayers. Furthermore, rhVim blocked neutrophil adhesion to P-selectin–coated surfaces. Binding assays showed that rhVim binds specifically to P-selectin and not to its counterreceptor, PSGL-1. Finally, in an endotoxin model of ALI in C57BL/6J mice, treatment with rhVim significantly decreased histologic findings of ALI. These data suggest a potential role for rhVim in attenuating inflammation through blocking P-selectin–PSGL-1 interactions.

No Child Left Behind: Liver Transplantation in Critically Ill Children

BACKGROUND Advances in critical care prolong survival in children with liver failure, allowing more critically ill children to undergo orthotopic liver transplantation (OLT). In order to justify the use of a scarce donor resource and avoid futile transplants, we sought to determine survival in children who undergo OLT while receiving pre-OLT critical care. STUDY DESIGN We analyzed 13,723 pediatric OLTs using the United Network for Organ Sharing (UNOS) database from 1987 to 2015, including 6,746 recipients in the Model for End-Stage Liver Disease/Pediatric End-Stage Liver Disease (MELD/PELD) era (2002 to 2015). There were 1,816 recipients (26.9%) admitted to the ICU at the time of transplantation. We also analyzed 354 pediatric OLT recipients at our center from 2002 to 2015, one of the largest institutional experiences. Sixty-five recipients (18.3%) were admitted to the ICU at the time of transplantation. Kaplan-Meier, volume threshold, and multivariable analyses were performed. RESULTS Patient survival improved steadily over the study period, (66% 1-year survival in 1987 vs 92% in 2015; p < 0.001). Our institutional experience of ICU recipients in the MELD/PELD era had acceptable outcomes (87% 1-year survival), even among our sickest recipients with vasoactive medications, mechanical ventilation, dialysis, and molecular adsorbent recirculating system requirements. Volume analysis revealed inferior outcomes (hazard ratio [HR] 1.68; 95% CI 1.11 to 2.51) in low-volume centers (<5 annual cases). Identifiable risk factors (previous transplantation, elevated serum sodium, hemodialysis, mechanical ventilation, body weight < 6 kg, and low center volume) increased risk of mortality. CONCLUSIONS This analysis demonstrates that the use of advanced critical care in children and infants with liver failure is justified because OLT can be performed on the sickest children and acceptable outcomes achieved. It is an appropriate use of a scarce donor allograft in a child who would otherwise succumb to a terminal liver disease.

Risk factors for infection after pediatric lung transplantation: XXXX

Although infection is the leading cause of death in the first year following pediatric lung transplantation, there are limited data on risk factors for early infection. Sepsis remains under‐recognized and under‐reported in the early post‐operative period for lung transplant recipients (LTR). We evaluated the incidence of infection and sepsis, and identified risk factors for infection in the early post‐operative period in pediatric LTRs. A retrospective review of medical records of LTRs at a large quaternary‐care hospital from January 2009 to March 2016 was conducted. Microbiology results on days 0‐7 after transplant were obtained. Sepsis was defined using the 2005 International Pediatric Consensus Conferencecriteria. Risk factors included history of recipient and donor infection, history of multi‐drug resistant (MDR) infection, nutritional status, and surgical times. Among the 98 LTRs, there were 22 (22%) with post‐operative infection. Prolonged donor ischemic time ≥7 hours, cardiopulmonary bypass(CPB) time ≥340 minutes, history of MDR infection and diagnosis of cystic fibrosis were significantly associated with infection. With multivariable regression analysis, only prolonged donor ischemic time remained significant (OR 4.4, 95% CI: 1.34‐14.48). Further research is needed to determine whether processes to reduce donor ischemic time could result in decreased post‐transplant morbidity.