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Coronary artery bypass grafting (CABG) triggers a systemic inflammatory response that may contribute to adverse outcomes. Dendritic cells (DC) and monocytes are immunoregulatory cells potentially affected by CABG, contributing to an altered immune state. This study investigated changes in DC and monocyte responses in CABG patients at 5 time‐points: admission, peri‐operative, ICU, day 3 and day 5. Whole blood from 49 CABG patients was used in an ex vivo whole blood culture model to prospectively assess DC and monocyte responses. Lipopolysaccharide (LPS) was added in parallel to model responses to an infectious complication. Co‐stimulatory and adhesion molecule expression and intracellular mediator production was measured by flow cytometry. CABG modulated monocyte and DC responses. In addition, DC and monocytes were immunoparalysed, evidenced by failure of co‐stimulatory and adhesion molecules (eg HLA‐DR), and intracellular mediators (eg IL‐6) to respond to LPS stimulation. DC and monocyte modulation was associated with prolonged ICU length of stay and post‐operative atrial fibrillation. DC and monocyte cytokine production did not recover by day 5 post‐surgery. This study provides evidence that CABG modulates DC and monocyte responses. Using an ex vivo model to assess immune competency of CABG patients may help identify biomarkers to predict adverse outcomes.

The Surviving Sepsis Campaign (SCC) and the American College of Critical Care Medicine (ACCM) guidelines recommend blood transfusion in sepsis when the haemoglobin concentration drops below 7.0 g/dL and 10.0 g/dL respectively, while the World Health Organisation (WHO) guideline recommends transfusion in septic shock ‘if intravenous (IV) fluids do not maintain adequate circulation’, as a supportive measure of last resort. Volume expansion using crystalloid and colloid fluid boluses for haemodynamic resuscitation in severe illness/sepsis, has been associated with adverse outcomes in recent literature. However, the volume expansion effect(s) following blood transfusion for haemodynamic circulatory support, in severe illness remain unclear with most previous studies having focused on evaluating effects of either different RBC storage durations (short versus long duration) or haemoglobin thresholds (low versus high threshold) pre-transfusion. • We describe the protocol for a pre-clinical randomised controlled trial designed to examine haemodynamic effect(s) of early volume expansion using packed RBCs (PRBCs) transfusion (before any crystalloids or colloids) in a validated ovine-model of hyperdynamic endotoxaemic shock. • Additional exploration of mechanisms underlying any physiological, haemodynamic, haematological, immunologic and tissue specific-effects of blood transfusion will be undertaken including comparison of effects of short (≤5 days) versus long (≥30 days) storage duration of PRBCs prior to transfusion. Protocol name: RESUS transfusion protocol, Keywords: Blood transfusion, Sepsis, Endotoxaemic shock, Packed red blood cells (PRBCs), Guidelines, Haemoglobin threshold, Storage duration

Purpose Cold static storage (CSS) is the standard method for heart preservation during transplantation (HTx). However, CSS beyond 4 hours increases the risk of primary graft dysfunction (PGD). Hypothermic ex vivo perfusion (HEVP) of donor hearts allows oxygen delivery during preservation, and may facilitate extended donor preservation without increasing PGD risk. We compared post-HTx survival, systemic inflammation and cardiac function following donor heart preservation by CSS (2 hrs) versus HEVP (2 and 8 hrs). Methods Brain death was induced in donor sheep for 24 hrs. Donor hearts were preserved by a) CSS for 2 hrs (n=7), b) HEVP for 2 hrs (n=4), or c) HEVP for 8 hrs (n=4). Orthotopic HTx was performed in matched recipients. Recipients were weaned from cardiopulmonary bypass and monitored for 6 hrs. Recipient blood was collected and assayed for inflammatory cytokines and cardiac markers. Cardiac function was assessed by echocardiography. Results Six-hour survival was 71% following CSS, and 100% following 2 and 8 hrs HEVP, respectively. Recipients systemic interleukin-6 and 8 levels were reduced using HEVP vs CSS. Post-HTx haemodynamic function was no different between groups, but HEVP reduced the requirement for vasoactive support compared to CSS (2 hrs CSS: 1.57±0.7; 2 hrs HEVP: 0.35±0.09; 8 hrs HEVP: 0.35±0.05 mmHg−1). HEVP was associated with reduced post-HTx lactate (2 hrs CSS: 11.4±1.8; 2 hrs HEVP: 5.2±0.7; 8 hrs HEVP: 6.7±1.3 mmol/L), more stable base excess and physiological pH in blood. Post-HTx cardiac function was no different between groups. Cardiac troponin I levels were comparable between CSS vs. 8 hrs HEVP, but reduced with 2 hrs HEVP. Conclusion Preliminary data on donor heart preservation by HEVP shows promising outcomes in comparison to CSS. Heart preservation by HEVP can be extended up to 8 hours, without compromising post-HTx recipient survival. HEVP may assist in overcoming limitations in preservation time associated with HTx, without increasing PGD risk.

Purpose Brain death (BD) causes metabolic and energetic imbalances leading to cardiac dysfunction, and predisposes the donor heart to further injury following heart transplantation (HTx). The metabolic mechanisms required for myocardial energy production during BD and subsequent HTx are poorly understood. Our aim was to determine the myocardial metabolic profile and mitochondrial function following donor BD and HTx. Methods Donor BD in sheep was induced by inflation of a catheter placed through the skull (catheter placement, but no inflation for SHAM), followed by 24 hrs monitoring, and heart procurement (n=6/group, BD vs. SHAM). Additional donor hearts exposed to BD/SHAM were flushed with cold St Thomas cardioplegia, and stored via cold static storage (CSS) for ∼2 hrs. Following standard orthotopic HTx, recipients were weaned off bypass and monitored for ≤6 hrs prior to heart procurement (n=4/group, BD-Tx vs. Sh-Tx). Cardiac mitochondrial function was assessed using high resolution respirometry. Metabolic profiles were determined in hearts using metabolomics. Cardiac mitochondrial function was also determined in two sheep that underwent HTx following BD and 8 hr hypothermic ex vivo perfusion (HEVP) preservation. Results BD caused significant right ventricular (RV) mitochondrial uncoupling (vs. SHAM). HTx following CSS also impaired RV mitochondrial function, with these effects more pronounced in hearts exposed to both donor BD and HTx. Early findings show that HEVP improved cardiac mitochondrial function post-HTx (vs. CSS). Metabolically, BD increased myocardial amino-acid utilisation and accumulation of glucose metabolites. Post-HTx, particularly in those exposed to donor BD, there was a significant decrease in metabolites involved in mitochondrial respiration (eg. NAD, Acetyl-CoA) and accumulation of fatty acids and xanthine (purine breakdown). Conclusion BD appears to trigger cardiac mitochondrial uncoupling. This may be a protective mechanism against higher amino-acid utilisation and glucose accumulation, in order to maintain adequate mitochondrial function for cell survival. HTx following CSS, particularly from BD donors, induces significant mitochondrial dysfunction, which occurs in response to upstream metabolic impairments. Strategies that improve cardiac mitochondrial function or metabolism (eg. HEVP) may assist to improve HTx outcomes.

Introduction: Packed red blood cell (PRBC) transfusion in septic shock is generally reserved as a last resort hemodynamic resuscitative measure or for haemoglobin less than 7.0 g/dL. Ex-vivo preservation and storage predispose PRBCs to biochemical / mechanical changes whose impact in a septic host is unknown. This randomised pre-clinical trial specifically evaluates select markers of cardiovascular stress in an ovine model of endotoxemic shock receiving either fresh or stored blood. Methods: Twenty six anesthetised and mechanically ventilated merino ewes were randomly divided into endotoxemic shock (n = 16) or healthy (n = 10) groups. Endotoxemic shock was induced over 4 hours by infusing an escalating dose (total 11.25 mcg / kg) of lipopolysaccharide, LPS ( E.coli 055:B5 ). During the last hour of LPS infusion, 10 mls / kg of either fresh, ≤ 5 days (n = 8) or stored, ≥ 30 days (n = 8) ovine packed PRBCs were administered. Healthy ewes received either fresh, ≤ 5 days (n = 5) or stored, ≥ 30 days (n = 5) ovine PRBCs at corresponding time points. Invasive haemodynamic monitoring and serial echocardiographic assessments were performed in all. Blood samples were measured for cardiac troponin I (cTnI), atrial natriuretic peptide (ANP) and hyaluronan at baseline (T0), 1, 3, 4, 5, 10 and 16h after which the ewes were euthanized. Results: Stored versus fresh PRBCs produced raised hyaluronan levels (P < 0.001) from T3 onwards in both sepsis and healthy groups and resulted in significant (P < 0.001) adverse haemodynamic changes (cardiac index, central venous pressure, pulmonary artery pressure and mixed venous oxygen saturation). Elevated ANP was observed between T3 to T10 (P < 0.001) in septic compared to healthy ewes but there was no association with age of PRBCs. No statistical differences were observed between groups for TnI. Conclusions: Sepsis and storage duration of PRBCs are associated with increased cardiovascular stress suggesting that stored PRBCs may predispose patients with septic shock to poor clinical outcomes.