Diaspirin Cross-linked Hemoglobin Research Articles

Orphan designation: Diaspirin cross-linked haemoglobin, Treatment of oesophageal cancer

Orphan designation: Diaspirin cross-linked haemoglobin, Treatment of oesophageal cancer

Orphan designation: Diaspirin cross-linked haemoglobin, Treatment of hepatocellular carcinoma

Orphan designation: Diaspirin cross-linked haemoglobin, Treatment of hepatocellular carcinoma

A Systematic Review and International Web-Based Survey of Randomized Controlled Trials in the Perioperative and Critical Care Setting: Interventions Increasing Mortality

Reducing mortality is a key target in critical care and perioperative medicine. The authors aimed to identify all nonsurgical interventions (drugs, techniques, strategies) shown by randomized trials to increase mortality in these clinical settings. A systematic review of the literature followed by a consensus-based voting process. A web-based international consensus conference. Two hundred fifty-one physicians from 46 countries. The authors performed a systematic literature search and identified all randomized controlled trials (RCTs) showing a significant increase in unadjusted landmark mortality among surgical or critically ill patients. The authors reviewed such studies during a meeting by a core group of experts. Studies selected after such review advanced to web-based voting by clinicians in relation to agreement, clinical practice, and willingness to include each intervention in international guidelines. The authors selected 12 RCTs dealing with 12 interventions increasing mortality: diaspirin-crosslinked hemoglobin (92% of agreement among web voters), overfeeding, nitric oxide synthase inhibitor in septic shock, human growth hormone, thyroxin in acute kidney injury, intravenous salbutamol in acute respiratory distress syndrome, plasma-derived protein C concentrate, aprotinin in high-risk cardiac surgery, cysteine prodrug, hypothermia in meningitis, methylprednisolone in traumatic brain injury, and albumin in traumatic brain injury (72% of agreement). Overall, a high consistency (ranging from 80% to 90%) between agreement and clinical practice was observed. The authors identified 12 clinical interventions showing increased mortality supported by randomized controlled trials with nonconflicting evidence, and wide agreement upon clinicians on a global scale.

Manipulating hemoglobin oxygenation using silica nanoparticles: a novel prospect for artificial oxygen carriers

AbstractRecently, nanoparticles have attracted much attention as new scaffolds for hemoglobin-based oxygen carriers (HBOCs). Indeed, the development of bionanotechnology paves the way for the rational design of blood substitutes, providing that the interaction between the nanoparticles and hemoglobin at a molecular scale and its effect on the oxygenation properties of hemoglobin are finely controlled. Here, we show that human hemoglobin has a high affinity for silica nanoparticles, leading to the adsorption of hemoglobin tetramers on the surface. The adsorption process results in a remarkable retaining of the oxygenation properties of human adult hemoglobin and sickle cell hemoglobin, associated with an increase of the oxygen affinity. The cooperative oxygen binding exhibited by adsorbed hemoglobin and the comparison with the oxygenation properties of diaspirin cross-linked hemoglobin confirmed the preservation of the tetrameric structure of hemoglobin loaded on silica nanoparticles. Our results show that silica nanoparticles can act as an effector for human native and mutant hemoglobin. Manipulating hemoglobin oxygenation using nanoparticles opens the way to the design of novel HBOCs.

Trials in adult critical care that show increased mortality of the new intervention: Inevitable or preventable mishaps?

Several promising therapies assessed in the adult critically ill in large, multicenter randomized controlled trials (RCTs) were associated with significantly increased mortality in the intervention arms. Our hypothesis was that there would be wide ranges in sponsorship (industry or not), type(s) of intervention(s), use of DSMBs, presence of interim analyses and early stopping rules, absolute risk increase (ARI), and whether or not adequate prior proof-of-principle Phase II studies were done of RCTs that found increased mortality rates of the intervention compared to control groups. We reviewed RCTs that showed a statistically significant increased mortality rate in the intervention compared to control group(s). We recorded source of sponsorship, sample sizes, types of interventions, mortality rates, ARI (as well as odds ratios, relative risks and number needed to harm), whether there were pre-specified interim analyses and early stopping rules, and whether or not there were prior proof-of-principle (also known as Phase II) RCTs. Ten RCTs (four industry sponsored) of many interventions (high oxygen delivery, diaspirin cross-linked hemoglobin, growth hormone, methylprednisolone, hetastarch, high-frequency oscillation ventilation, intensive insulin, NOS inhibition, and beta-2 adrenergic agonist, TNF-α receptor) included 19,126 patients and were associated with wide ranges of intervention versus control group mortality rates (25.7–59 %, mean 29.9 vs 17–49 %, mean 25 %, respectively) yielding ARIs of 2.6–29 % (mean 5 %). All but two RCTs had pre-specified interim analyses, and seven RCTs were stopped early. All RCTs were preceded by published proof-of-principle RCT(s), two by the same group. Seven interventions (except diaspirin cross-linked hemoglobin and the NOS inhibitor) were available for use clinically at the time of the pivotal RCT. Common, clinically available interventions used in the critically ill were associated with increased mortality in large, pivotal RCTs even though safety was often addressed by interim analyses and early stopping rules.

Polymerization of modified diaspirin cross-linked hemoglobin (DCLHb) with 1,6-bismaleimic-hexane

Increasing the size of hemoglobin (Hb) by polymerization offers the benefits of reduced renal clearance and increased duration in the vascular circulation. With this goal, diaspirin cross-linked hemoglobin (DCLHb) was modified in order to keep one thiol group on the surface and then polymerized with 1,6-bismaleimic-hexane (1,6-BMH) to increase the molecular weight. The HPLC results indicated that approximate 20% dimers to tetramers of DCLHb desired were achieved after the polymerization. It was also demonstrated that the oxygen-carrying capacity of the products was similar to natural heme. The present study is expected to improve the efficacy of the DCLHb as an oxygen therapeutic agent.

Emergency Resuscitation of Patients Enrolled in the US Diaspirin Cross-linked Hemoglobin (DCLHb) Clinical Efficacy Trial.

Optimal emergent management of traumatic hemorrhagic shock patients requires a better understanding of treatment provided in the prehospital/Emergency Medical Services (EMS) and emergency department (ED) settings. Hypothesis/Problem Described in this research are the initial clinical status, airway management, fluid and blood infusions, and time course of severely-injured hemorrhagic shock patients in the EMS and ED settings from the diaspirin cross-linked hemoglobin (DCLHb) clinical trial. Data were analyzed from 17 US trauma centers gathered during a randomized, controlled, single-blinded efficacy trial of a hemoglobin solution (DCLHb) as add-on therapy versus standard therapy. Among the 98 randomized patients, the mean EMS Glasgow Coma Scale (GCS) was 10.6 (SD = 5.0), the mean EMS revised trauma score (RTS) was 6.3 (SD = 1.9), and the mean injury severity score (ISS) was 31 (SD = 17). Upon arrival to the ED, the GCS was 20% lower (7.8 (SD = 5.3) vs 9.7 (SD = 6.3)) and the RTS was 12% lower (5.3 (SD = 2.0) vs 6.0 (SD = 2.1)) than EMS values in blunt trauma patients (P < .001). By ED disposition, 80% of patients (78/98) were intubated. Rapid sequence intubation (RSI) was utilized in 77% (60/78), most often utilizing succinylcholine (65%) and midazolam (50%). The mean crystalloid volume infused was 4.2 L (SD = 3.4 L), 80% of which was infused within the ED. Emergency department blood transfusion occurred in 62% of patients, with an average transfused volume of 1.2 L (SD = 2.0 L). Blunt trauma patients received 2.1 times more total fluids (7.4 L vs 3.5 L, < .001) and 2.4 times more blood (2.4 L vs 1.0 L, P < .001). The mean time of patients taken from injury site to operating room (OR) was 113 minutes (SD = 87 minutes). Twenty-one (30%) of the 70 patients taken to the OR from the ED were sent within 60 minutes of the estimated injury time. Penetrating trauma patients were taken to the OR 52% sooner than blunt trauma patients (72 minutes vs 149 minutes, P < .001). Both GCS and RTS decreased prior to ED arrival in blunt trauma patients. Intubation was performed using RSI, and crystalloid infusion of three times the estimated blood loss volume (L) and blood transfusion of the estimated blood loss volume (L) were provided in the EMS and ED settings. Surgical intervention for these trauma patients most often occurred more than one hour from the time of injury. Penetrating trauma patients received surgical intervention more rapidly than those with a blunt trauma mechanism.

Shock index and prediction of traumatic hemorrhagic shock 28-day mortality: data from the DCLHb resuscitation clinical trials.

IntroductionTo assess the ability of the shock index (SI) to predict 28-day mortality in traumatic hemorrhagic shock patients treated in the diaspirin cross-linked hemoglobin (DCLHb) resuscitation clinical trials.MethodsWe used data from two parallel DCLHb traumatic hemorrhagic shock efficacy trials, one in U.S. emergency departments, and one in the European Union prehospital setting to assess the relationship between SI values and 28-day mortality.ResultsIn the 219 patients, the mean age was 37 years, 64% sustained a blunt injury, 48% received DCLHb, 36% died, and 88% had an SI≥1.0 at study entry. The percentage of patients with an SI≥1.0 dropped by 57% (88 to 38%) from the time of study entry to 120 minutes after study resuscitation (p<0.001). Patients with a SI≥1.0, 1.4, and 1.8 at any time point were 2.3, 2.7, and 3.1 times, respectively, more likely to die by 28 days than were patients with SI values below these cutoffs (p<0.001). Similarly, after 120 minutes of resuscitation, patients with a SI≥1.0 were 3.9× times more likely to die by 28 days (40 vs. 15%, p<0.001). Although the distribution of SI values differed based on treatment group, the receiver operator characeristics data showed no difference in SI predictive ability for 28-day mortality in patients treated with DCLHb.ConclusionIn these traumatic hemorrhagic shock patients, the shock index correlates with 28-day mortality, with higher SI values indicating greater mortality risk. Although DCLHb treatment did alter the distribution of SI values, it did not influence the ability of the SI to predict 28-day mortality.

The Use of the Revised Trauma Score as an Entry Criterion in Traumatic Hemorrhagic Shock Studies: Data from the DCLHb Clinical Trials

The Revised Trauma Score (RTS) has been proposed as an entry criterion to identify patients with mid-range survival probability for traumatic hemorrhagic shock studies. Determination of which of four RTS strata (1-3.99, 2-4.99, 1-4.99, and 2-5.99) identifies patients with predicted and actual mortality rates near 50% for use as an entry criterion in traumatic hemorrhagic shock clinical trials. Existing database analysis in which demographic and injury severity data from two prior international Diaspirin Cross-Linked Hemoglobin (DCLHb) clinical trials were used to identify an RTS range that could be an optimal entry criterion in order to find the population of trauma patients with mid-range predicted and actual mortality rates. Of 208 study patients, the mean age was 37 years, 65% sustained blunt trauma, 49% received DCLHb, and 57% came from the European Union study arm. The mean values were: ISS, 31 (SD = 18); RTS, 5.6 (SD = 1.8); and Glasgow Coma Scale (GCS), 10.4 (SD = 4.8). The mean TRISS-predicted mortality was 34% and the actual 28-day mortality was 35%. The initially proposed 1-3.99 RTS range (n = 41) had the highest predicted (79%) and actual (71%) mortality rates. The 2-5.99 RTS range (n = 79) had a 62% predicted and 53% actual mortality, and included 76% blunt trauma patients. Removal of GCS <5 patients from this RTS 2-5.99 subgroup caused a 48% further reduction in eligible patients, leaving 41 patients (20% of 208 total patients), 66% of whom sustained a blunt trauma injury. This subgroup had 54% predicted and 49% actual mortality rates. Receiver operator curve (ROC) analysis found the GCS to be as predictive of mortality as the RTS, both in the total patient population and in the RTS 2-5.99 subgroup. The use of an RTS 2-5.99 inclusion criterion range identifies a traumatic hemorrhagic shock patient subgroup with predicted and actual mortality that approach the desired 50% rate. The exclusion of GCS <5 from this RTS 2-5.99 subgroup patients yields a smaller, more uniform patient subgroup whose mortality is more likely related to hemorrhagic shock than traumatic brain injury. Future studies should examine whether the RTS or other physiologic criteria such as the GCS score are most useful as traumatic hemorrhagic shock study entry criteria.

Hemoglobin-based Oxygen Carriers for Reversing Hypotension and Shock

Cardiothoracic Anesthesiology and Critical Care, Emory University School of Medicine, Atlanta, Georgia. jlevy01@emory.eduEFFORTS to develop a blood substitute, especially for therapy of hemorrhagic shock, have been ongoing. Most clinical trials have focused on hemoglobin-based oxygen carriers (HBOCs) rather than perfluorochemical derivatives. Although HBOCs are purified, synthesized, or modified hemoglobins from human, bovine, or recombinant sources, they most often originate from human senescent blood.1,2Traditional thinking supports the idea that HBOCs may facilitate volume expansion and stabilize shock patients until erythrocytes become available. Unfortunately, it is not that simple.Hemoglobin is an oxygen-carrying transport molecule physiologically contained in erythrocytes and evolutionarily designed as such. However, outside erythrocytes, the molecules function quite differently, with an altered affinity for oxygen in a different milieu of the circulation.3,4Unmodified hemoglobin solutions have different oxygen affinities when outside red cells; the chains have much higher oxygen affinity than do native or cross-linked hemoglobins.3,4As a result, purified hemoglobins have been modified by polymerization, cross-linking, or pegylation to improve oxygen delivery, increase their half-life, and alter their safety profile. The oncotic pressure exerted by these solutions varies widely. Each HBOC has been uniquely modified to improve its oxygen-carrying and -delivery capabilities and contains different amounts of hemoglobin as dosed. However, despite these physiochemical modifications, most HBOCs significantly scavenge nitric oxide (NO). This process is thought to account for many of the adverse effects and outcomes associated with these agents.1HBOCs were never intended to have this property. Their use for resuscitation or intraoperative treatment was based on their volume replacement and oxygen-carrying capacity, which is the basis of the current study in Anesthesiology.5In the endothelium, throughout the vascular arterial bed, NO is important in maintaining vascular tone. Under physiologic conditions and within the erythrocyte membrane, hemoglobin is mostly insulated from the vascular endothelium. However, once released, free hemoglobin and HBOCs are potent NO scavengers. Because of this mechanism, most HBOCs have been shown to cause systemic and pulmonary artery vasoconstriction, though an oxygenated polyethylene glycol–modified hemoglobin molecule (MP4OX [formerly known as Hemospan®]; Sangart, Inc., San Diego, CA) is thought to have enhanced NO synthesis (NOS) as a result of nitrite reductase properties because of β-chain cysteine pegylation in the 93 position.1,6–8Thus, one of the major drawbacks in studying HBOCs for the treatment of hemorrhagic shock is that they avidly bind NO, causing pulmonary and systemic vasoconstriction.8–10The ferric group of iron in hemoglobin is the primary mechanism by which NO is scavenged. This is also why, in part, inhaled NO as used in clinical concentrations has pulmonary artery vasodilating specificity with minimal effects on the systemic vasculature: scavenging occurs via erythrocyte hemoglobin.11HBOCs increase pulmonary and systemic blood pressures, that may, in turn, impair autoregulation of vascular flow in organ systems, as demonstrated in animal models. This process may account for adverse events seen clinically with HBOCs.2Furthermore, endothelial dysfunction may enhance vasoconstriction, as in the case of patients with atherosclerotic vascular disease.12In Anesthesiology this month, Olofsson et al .5report a clinical study of MP4OX as a low-volume, oxygen-therapeutic agent that is intended to deliver oxygen to ischemic tissue.5The authors used this oncotically active agent to prevent hypotensive episodes and also to determine its safety in a surgical setting. Olofsson et al .5observed 367 patients from 18 sites in six countries during elective primary hip arthroplasty with spinal anesthesia. Patients were randomized to receive MP4OX or hydroxyethyl starch (HES130/0.4) at induction of spinal anesthesia. A second dose was made available if a predefined drop in systolic blood pressure occurred.The proportion of patients with one or more hypotensive episode(s), the primary endpoint of the study, was significantly lower in the MP4OX group (66.1%) versus HES 130/0.4 controls (90.2%). However, when compared with controls, more MP4OX-treated patients experienced adverse events (72.7 vs. 61.4%), including nausea (20.8 vs. 12.0%) and hypertension (4.4 vs. 0.5%). None of the hypertensive adverse events observed occurred during the operative and 6-h postoperative recovery period.Another consistent finding in HBOC studies has been transient elevations in certain laboratory assays, including liver function tests and troponin.13Such was the case also in this study by Olofsson et al .5Despite this finding, however, there were no differences between study groups with regard to morbidity or serious adverse events. The authors5conclude that the adverse event profile does not support the use of MP4OX in routine, low-risk surgical patients.Reports from an earlier phase 2 clinical trial of MP4OX enrolled patients undergoing orthopedic surgical procedures.14In that investigation,14patients were randomized to receive MP4OX or Ringer's acetate (30 patients per group). However, the earlier investigation by Olofsson et al .14was a safety study. Olofsson et al .5report that MP4OX mildly elevates hepatic enzymes and lipase and was associated with less hypotension and more bradycardia events.There are ongoing controversies regarding the safety of HBOCs in clinical trials. One analysis in particular was criticized for including different therapeutic areas and HBOCs in the analysis given the inherent variability of the patients studied and differences between HBOCs.9Previous clinical trials by Baxter International, Inc. (Deerfield, IL) and Northfield Laboratories (Evanston, IL) have tested the use of HBOCs in trauma.2,14However, because hemorrhage and shock are the leading cause of civilian and battlefield mortality, there remain, in my view, potential applications of HBOCs in treating such injuries, as noted by the S. Armed Forces Blood Program.15In 2009, the results of a National Institutes of Health and U.S. Food and Drug Administration workshop were published regarding HBOC safety, addressing current status and future direction of these agents.2,14Multiple HBOCs are available and have been evaluated in numerous clinical studies, as discussed. My experience with HBOCs was primarily with Hemopure® (Biopure Corporation, Cambridge, MA), a cross-linked hemoglobin polymer from bovine blood.16,17I also wrote an editorial for Anesthesiology about diaspirin cross-linked hemoglobin, DCLHb (HemAssist; Baxter Healthcare Corporation, Deerfield, IL), as studied in clinical trials for coronary artery surgery.7,18However, adverse effects detected in other studies ended clinical investigations for this agent.7NO scavenging has consistently been held responsible for many of the adverse effects and events observed with HBOCs, including myocardial injury and infarction, gastrointestinal symptoms (e.g ., nausea), and ischemic injury in shock.2,14Heme itself may be toxic. In addition, MP4OX used in low volumes and having a reduced hemoglobin concentration may account for the low rate of reported adverse effects. It is noteworthy that the current Olofsson et al .5study reported adverse effects similar to those detailed in the workshop report by the National Institutes of Health and U.S. Food and Drug Administration.2Clinical studies have also investigated agents that inhibit NOS, specifically the enzyme involved in NO production. These investigations report conflicting results, documenting beneficial and detrimental effects.19Because the precursor for NO is arginine, most NOS inhibitors are derivatives of arginine. A study in humans evaluated L -n-monomethyl-arginine, an NOS inhibitor, in 79 patients after acute myocardial infarction and cardiogenic shock, despite an open infarct-related artery.20Using incremental dosing, mean change in mean arterial pressure ranged from 4.8 to 11.6 mmHg at 15 min, but not at 2 h. However, cardiogenic shock is probably not a great model when evaluating NO inhibition. Using an in vitro human vasodilation model, we reported the effects of the NOS inhibitors methylene blue and n(G)-monomethyl-L-arginine, as well as prostaglandin inhibitors (indometacin).21We noted that inhibition of either NOS or prostaglandin pathways did not completely reverse vasodilation; both effects were required for mediator-induced vasodilation.21Thus, in severe vasodilatory shock, including anaphylactic shock, therapeutic approaches must be multimodal and target more than NO.In summary, it is unlikely that we will have a NO-scavenging agent to treat hypotension or shock until the molecule is sufficiently safe for any benefits provided. However, Olofsson et al .5(and Sangart, Inc.) are to be congratulated on this new data—and for safety findings and information that may help future researchers who wish to investigate novel oxygen-therapeutic molecules for other, more appropriate clinical indications. The paradigm of using a molecule that scavenges NO as an adverse effect for a primary therapeutic endpoint is important. It may provide a useful path for future studies in this therapeutic area.Cardiothoracic Anesthesiology and Critical Care, Emory University School of Medicine, Atlanta, Georgia. jlevy01@emory.edu

Vasoactive drugs for acute stroke.

It is unclear whether blood pressure (BP) should be altered actively during the acute phase of stroke. To assess the effect of lowering or elevating BP in people with acute stroke, and the effect of different vasoactive drugs on BP in acute stroke. We searched the Cochrane Stroke Group Trials Register (last searched June 2009), the Cochrane Central Register of Controlled Trials (CENTRAL) (The Cochrane Library Issue 4, 2009), MEDLINE (1966 to October 2009), EMBASE (1980 to October 2009), and Science Citation Index (1981 to October 2009). Randomised trials of interventions that would be expected, on pharmacological grounds, to alter BP in patients within one week of the onset of acute stroke. Two review authors independently applied the trial inclusion criteria, assessed trial quality, and extracted data. We identified 131 trials involving in excess of 18,000 patients; a further 13 trials are ongoing. We obtained data for 43 trials (7649 patients). Among BP-lowering trials, beta receptor antagonists lowered BP (early systolic BP (SBP) mean difference (MD) -6.1 mmHg, 95% CI -11.4 to -0.9; late SBP MD -4.9 mmHg, 95% CI -10.2 to 0.4; late diastolic BP (DBP) MD -4.5 mmHg, 95% CI -7.8 to -1.2). Oral calcium channel blockers (CCB) lowered BP (late SBP MD -3.2 mmHg, 95% CI -5.4 to -1.1; early DBP MD -2.5, 95% CI -5.6 to 0.7; late DBP MD -2.1, 95% CI -3.5 to -0.7). Nitric oxide donors lowered BP (early SBP MD -10.3 mmHg, 95% CI -17.6 to -3.0). Prostacyclin lowered BP (late SBP MD, -7.7 mmHg, 95% CI -15.6 to 0.2; late DBP MD -3.9 mmHg, 95% CI -8.1 to 0.4). Among BP-increasing trials, diaspirin cross-linked haemoglobin (DCLHb) increased BP (early SBP MD 15.3 mmHg, 95% CI 4.0 to 26.6; late SBP MD 15.9 mmHg, 95% CI 1.8 to 30.0). None of the drug classes significantly altered outcome apart from DCLHb which increased combined death or dependency (odds ratio (OR) 5.41, 95% CI 1.87 to 15.64). There is not enough evidence to evaluate reliably the effect of altering BP on outcome after acute stroke. However, treatment with DCLHb was associated with poor clinical outcomes. Beta receptor antagonists, CCBs, nitric oxide, and prostacyclin each lowered BP during the acute phase of stroke. In contrast, DCLHb increased BP.

Diaspirin Cross-Linked Hemoglobin Infusion Did Not Influence Base Deficit and Lactic Acid Levels in Two Clinical Trials of Traumatic Hemorrhagic Shock Patient Resuscitation

Diaspirin cross-linked hemoglobin (DCLHb) has demonstrated a pressor effect that could adversely affect traumatic hemorrhagic shock patients through diminished perfusion to vital organs, causing base deficit (BD) and lactate abnormalities. Data from two parallel, multicenter traumatic hemorrhagic shock clinical trials from 17 US Emergency Departments and 27 European Union prehospital services using DCLHb, a hemoglobin-based resuscitation fluid. In the 219 patients, the mean age was 37.3 years, 64% of the patients sustained a blunt injury, 48% received DCLHb resuscitation, and the overall 28-day mortality rate was 36.5%. BD data did not differ by treatment group (DCLHb vs. normal saline [NS]) at any time point. Study entry BD was higher in patients who died when compared with survivors in both studies (US: -14.7 vs. -9.3 and European Union: -11.1 vs. -4.1 mEq/L, p < 0.003) and at the first three time points after resuscitation. No differences in BD based on treatment group were observed in either those who survived or those who died from the hemorrhagic shock. US lactate data did not differ by treatment group (DCLHb vs. NS) at any time point. Study entry lactates were higher in US patients who ultimately died when compared with survivors (82.4 vs. 56.1 mmol/L, p < 0.003) and at all five postresuscitation time points. No lactate differences were observed between DCLHb and NS survivors or in those who died based on treatment group. Although patients who died had more greatly altered perfusion than those who survived, DCLHb treatment of traumatic hemorrhagic shock patients was not associated with BD or lactate abnormalities that would indicate poor perfusion.

Pharmacokinetics of diaspirin cross-linked haemoglobin in a rat model of hepatic cirrhosis.

The aim of the study was to evaluate the effect of cirrhosis on the disposition of the haemoglobin-based oxygen carrier, diaspirin cross-linked haemoglobin (DCLHb). Cirrhosis was induced in male Sprague-Dawley rats (200-250 g) by inhalational exposure to carbon tetrachloride (CCl4), over a period of 6 weeks. Pharmacokinetic evaluation was performed after a single intravenous bolus administration of DCLHb (400 mg kg(-1)). Serum biochemistry, including aspartate transaminase, alkaline phosphatase, bile acids, serum albumin, and serum creatinine, were measured in CCl4-treated (n = 6) and age-matched control (n = 6) rats. After 6 weeks, the jugular vein and carotid artery were cannulated for bolus DCLHb administration (400 mg kg(-1)) and blood sampling, respectively, in both groups of rats. Cirrhosis produced significant (P < 0.05) elevations in alkaline phosphatase (497.4 +/- 84.8 U L(-1) vs 241.2 +/- 5.1 U L(-1)), aspartate transaminase (920.5 +/- 190.9 U L(-1) vs 238.2 +/- 118.1 U L(-1)) and bile acids (333.8 +/- 77.3 mg dL(-1) vs 43.8 +/- 4.2 mg dL(-1)) compared with the control group. No significant renal dysfunction was observed as a result of CCl4 exposure. Plasma DCLHb concentrations declined approximately log-linearly. Systemic clearance of DCLHb was estimated to be 2.2 +/- 0.7 mL h(-1) in the treatment group and was slightly, but not significantly, less in the control group (3.6 +/- 1.7 mL h(-1)). There was also a trend toward a longer elimination half-life in the treatment group (4.7 +/- 2.2 h) compared with the control group (3.8 +/- 0.8 h), although this difference was not statistically significant. Cirrhosis does not significantly alter the disposition of DCLHb perhaps due to increased extra-hepatic metabolism by the reticulo-endothelial system.

THE LACK OF CONSISTENT DIASPIRIN CROSS-LINKED HEMOGLOBIN INFUSION BLOOD PRESSURE EFFECTS IN THE US AND EU TRAUMATIC HEMORRHAGIC SHOCK CLINICAL TRIALS

Hemoglobin solutions have demonstrated a pressor effect that could adversely affect hemorrhagic shock patient resuscitation through accelerated hemorrhage, diminished perfusion, or inadequate resuscitation. Data from two parallel, multicenter traumatic hemorrhagic shock clinical trials in 17 US emergency departments and in 27 EU prehospital systems using diaspirin cross-linked hemoglobin (DCLHb), a hemoglobin-based resuscitation fluid. In the 219 patients, patients were 37 years old, 64% sustained blunt injury, 48% received DCLHb, and 36% expired. Although mean systolic blood pressure (SBP) and diastolic blood pressure values differed at 2 of the 10 measured time points, blood pressure (BP) curve analysis showed no SBP, diastolic blood pressure, or MAP differences based on treatment. Although SBP values 160 and 120 mmHg or greater were 2.2x and 2.6x more frequently noted in survivors, they were not more common with DCLHb use or in DCLHb patients who expired in US study nonsurvivors or in any EU study patients. Systolic blood pressure values 160 and 120 mmHg or greater were 2.8x and 1.3x more frequently noted in DCLHb survivors as compared with normal saline survivors. Only 3% of the BP variation noted could be attributed to DCLHb use, and as expected, injury severity and baseline physiologic status were stronger predictors. In the United States alone, treatment group was not correlated by regression with BP at any time point. Neither mean BP readings nor elevated BP readings were correlated with DCLHb treatment of traumatic hemorrhagic shock patients. As such, no clinically demonstrable DCLHb pressor effect could be directly related to the adverse mortality outcome observed in the US study.

New phosphorescence quenching oxygen measurements technique yields unusual tissue and plasma Po2 distributions

LETTERS TO THE EDITORNew phosphorescence quenching oxygen measurements technique yields unusual tissue and plasma Po2 distributionsAmy G. Tsai, Pedro Cabrales, Paul C. Johnson, and Marcos IntagliettaAmy G. Tsai, Pedro Cabrales, Paul C. Johnson, and Marcos IntagliettaPublished Online:01 May 2007https://doi.org/10.1152/japplphysiol.00122.2007MoreSectionsPDF (60 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInEmailWeChat To the Editor: Wilson et al. (9) report using a novel group of chromophores for measuring oxygen pressure in the interstitial space and blood plasma, proposing that the histogram of Po2 volume distribution measured in the thigh muscle of mice can be used to 1) describe the partition of tissue and intravascular Po2 and 2) characterize the difference between intra- and extravascular Po2 or the vessel wall gradient. Both contentions would appear to be at odds with the results and data presented.Their histograms (Fig. 2) show intra- and extravascular Po2 volume fractions of ∼10 and 20%, respectively, with Po2 values of 100–140 Torr. The maximum Po2 in arterial blood leaving the lungs is ∼100 mmHg, and the existence of a longitudinal gradient downstream is a well established feature of Po2 in the vasculature (8). Even in small animals, it is not clear how the Po2 of blood in the vasculature or the tissue of the limbs can exceed Po2 of blood in the lungs. A measurement technique that records such high Po2 values in the interstitial and the intravascular space raises questions about how the measurements relate to the distribution of Po2 in the tissue reported by previous investigators.The review of Tsai et al. (8) on oxygen gradients in the microcirculation shows that the highest Po2 value recorded in 12 studies of arterioles up to 100 μm using either the microelectrode or the phosphorescence technique was ∼80 Torr. Boeghhold and Johnson (1), 1988, recorded in 55-μm-diameter arterioles of the cat sartorious muscle a periarteriolar Po2 of 52 ± 3 mmHg (SE) when tissue Po2 was 23 ± 3 mmHg (SE) using Whalen microelectrodes. Smith et al. (6), 2004, in similar- sized arterioles in rat spinotrapezius muscle, measured a Po2 of 37 ± 2 mmHg and 31 ± 3 mmHg in venules with the oxygen phosphorescence quenching technique. The review reports the highest venular Po2 found in eight studies was ∼40 mmHg (8). Thus the finding of Wilson et al. (9) that 30% of the intravascular volume had Po2 values over 80 Torr (their Fig. 2) is difficult to reconcile with previous reports. Regarding tissue Po2, Nolte et al. (4), 1997, used the multiwire surface electrodes to map tissue Po2 in the awake hamster window chamber model and reported tissue Po2 values of 19 ± 2 mmHg, showing no values >40 mmHg. Wilson et al. found that >60% of their tissue values were above that level. These large discrepancies with the findings of Wilson et al. call into question conclusions regarding a gradient between intravascular and tissue measurements on the basis of their technique.The problem with the tissue Po2 measurements by Wilson et al. (9) may be due in part to their methodology, which requires direct injection of a chromophore into the tissue. They inserted a needle 1 cm into the tissue, delivering injections of 20 μl at three neighboring locations. This procedure is very likely to damage the microcirculatory vessels with leakage of blood into the tissue, elevation of local tissue pressure, and blood flow stoppage. Thus the “tissue” values will be compromised by red blood cells and plasma in the area of measurements. Assuming that the mouse tissue involved was 1 cm3 in volume, introducing 60 μl caused at least a 6% tissue volume expansion, potentially increasing tissue fluid by ∼50%, a major perturbation in the tissue.The authors (9) compare the tissue Po2 histograms in the 5% volume fraction with the lowest Po2 (average 15.9 ± 6.1 Torr) to the same volume fraction tagged by the intravascular marker (average 17.4 ± 4.0 Torr), concluding that 1.5 Torr difference does not represent a capillary wall gradient “… supporting the hypothesis that the vessel walls do not provide a significant barrier to oxygen delivery.” In addition to the cited technique problems, no evidence is given to justify choosing the lowest 5% of the values from each compartment to represent capillaries and adjacent tissue. They indicate that the 1.5 Torr difference contradicts the finding of Tsai et al. (7), 1998, in arterioles while apparently recognizing that arterioles cannot be compared to capillaries. Their data were from awake animals, where “… oxygen histograms were highly variable, and many had very broad histograms with peaks at Po2 values of 60 Torr and above. These broad distributions with increased high Po2 values correlated with the visual evidence of agitation …” Their data show a gradient of 6.1 Torr for isofluorane-anesthetized animals (free of motion artifacts?) for the same histogram Po2 range. Although it is likely fortuitous, this is the gradient previously reported for hamster capillaries (3).Assigning a histogram Po2 level to a microvascular anatomical feature is problematic since the microvascular Po2 distribution is “U” shaped, many arterioles and venules having similar Po2 values. The lowest Po2 values may be due to signals from outside the capillary, measurement of tissue Po2 near collecting venules or from Po2 signals from terminal lymphatics, the lowest Po2 in the tissue (2). Venules contain most of the blood volume in the microcirculation, whose Po2 is above tissue Po2 (5). The large intravascular venular blood volume vs. capillary and arteriolar volume should bias Po2 toward lower values; however, the results of Wilson et al. (9) show that more than half of intravascular volume has Po2 >50 Torr.In summary, this new technique for measuring tissue and intravascular Po2 appears to be flawed since it reports tissue and intravascular Po2 measurements (100–140 Torr) not compatible with the physics of oxygen distribution in a mammal. Very high Po2 values in the distribution indicate the presence of effects that probably influence the whole data set. In the muscle microcirculation there is no evidence of Po2 values greater than ∼60–70 mmHg, and, as reported by previous investigators, most values are much lower. A careful reevaluation of the in vivo application of this method would appear to be in order.GRANTSThis research was supported by National Heart, Lung, and Blood Institute Grants HL-76182, HL-62318, HL-62354, and HL-64395.REFERENCES1 Boegehold MA, Johnson PC. Periarteriolar and tissue Po2 during sympathetic escape in skeletal muscle. Am J Physiol Heart Circ Physiol 254: H929–H936, 1988.Link | ISI | Google Scholar2 Hangai-Hoger N, Cabrales P, Briceno JC, Tsai AG, Intaglietta M. Microlymphatic and tissue oxygen tension in the rat mesentery. Am J Physiol Heart Circ Physiol 286: H878–H883, 2004.Link | ISI | Google Scholar3 Intaglietta M, Johnson PC, Winslow RM. Microvascular and tissue oxygen distribution. Cardiovasc Res 32: 632–643, 1996.Crossref | PubMed | ISI | Google Scholar4 Nolte D, Botzlar A, Pickelmann S, Bouskela E, Messmer K. Effects of diaspirin-cross-linked hemoglobin (DCLHb) on the microcirculation of striated skin muscle in the hamster: a study on safety and toxicity. J Lab Clin Med 130: 314–327, 1997.Crossref | PubMed | Google Scholar5 Saltzman DJ, Toth A, Tsai AG, Intaglietta M, Johnson PC. Oxygen tension distribution in postcapillary venules in resting skeletal muscle. Am J Physiol Heart Circ Physiol 285: H1980–H1985, 2003.Link | ISI | Google Scholar6 Smith LM, Barbee RW, Ward KR, Pittman RN. Prolonged tissue Po2 reduction after contraction in spinotrapezius muscle of spontaneously hypertensive rats. Am J Physiol Heart Circ Physiol 287: H401–H407, 2004.Link | ISI | Google Scholar7 Tsai AG, Friesenecker B, Mazzoni MC, Kerger H, Buerk DG, Johnson PC, Intaglietta M. Microvascular and tissue oxygen gradients in the rat mesentery. Proc Natl Acad Sci USA 95: 6590–6595, 1998.Crossref | PubMed | ISI | Google Scholar8 Tsai AG, Johnson PC, Intaglietta M. Oxygen gradients in the microcirculation. Physiol Rev 83: 933–963, 2003.Link | ISI | Google Scholar9 Wilson DF, Lee WM, Makonnen S, Finikova O, Apreleva S, Vinogradov SA. Oxygen pressures in the interstitial space and their relationship to those in the blood plasma in resting skeletal muscle. J Appl Physiol 101: 1648–1656, 2006.Link | ISI | Google ScholarAUTHOR NOTESAddress for reprint requests and other correspondence: M. Intaglietta, Univ. of California, San Diego, Dept. of Bioengineering, 9500 Gilman Dr., La Jolla, CA 92093–0412 (e-mail: [email protected]) Download PDF Previous Back to Top Next FiguresReferencesRelatedInformationCited ByEvaluation of multi-exponential curve fitting analysis of oxygen-quenched phosphorescence decay traces for recovering microvascular oxygen tension histograms3 November 2010 | Medical & Biological Engineering & Computing, Vol. 48, No. 12Monitoring oxygen uptake in 3D tissue engineering scaffolds by phosphorescence quenching microscopy22 April 2010 | Biotechnology Progress, Vol. 26, No. 5In Vivo Mitochondrial Oxygen Tension Measured by a Delayed Fluorescence Lifetime TechniqueBiophysical Journal, Vol. 95, No. 8METHODS FOR STUDYING THE PHYSIOLOGY OF KIDNEY OXYGENATIONClinical and Experimental Pharmacology and Physiology, Vol. 33Reply to Tsai, Cabrales, Johnson, and IntagliettaDavid F. Wilson, William M. F. Lee, Sosina Makonnen, Olga Finikova, Sofia Apreleva, and Sergei A. Vinogradov1 May 2007 | Journal of Applied Physiology, Vol. 102, No. 5 More from this issue > Volume 102Issue 5May 2007Pages 2081-2082 Copyright & PermissionsCopyright © 2007 the American Physiological Societyhttps://doi.org/10.1152/japplphysiol.00122.2007PubMed17483445History Published online 1 May 2007 Published in print 1 May 2007 Metrics

EFFECT OF PRETREATMENT WITH HEME-BASED SOLUTIONS ON LIVER MICROCIRCULATION AFTER HEMORRHAGIC SHOCK

Pretreatment with hemoglobin-based oxygen carriers (diaspirin-crosslinked hemoglobin, DCLHb; hemoglobin glutamer-200, HbG) induces heme oxygenase-1 (HO-1), but increases organ dysfunction and mortality in a rat model of hemorrhagic shock. In contrast, pretreatment with the non oxygen-carrying hemin arginate (HAR) reduces shock-induced organ failure and mortality probably through HO-1 induction. The NO-scavenging and vasoconstrictive effects of free hemoglobin may affect microcirculation after shock. Aim of the study was to assess the effect of pretreatment with DCLHb, HbG or HAR on liver microcirculation. 24 h after pretreatment with DCLHb (1 g/kg), HbG (1 g/kg) or HAR (5 mg/kg) rats were subjected to 1 h hemorrhage (MAP: 35 ± 5 mmHg)/2 h resuscitation. Liver microcirculation was ssessed by intravital microscopy. Animals treated with Ringer's solution served as controls (vehicle). DCLHb, HbG, and HAR induced HO-1 in the liver. However, DCLHb and HbG-pretreatment did not improve sinusoidal perfusion and increased NAD(P)H autofluorescence in liver tissue similar to vehicle-treated animals. In contrast, HAR-pretreatment significantly improved sinusoidal perfusion. Blockade of the HO-pathway by SnMP-IX (10 μmol/kg) after HAR-pretreatment abolished the protective effects. Although DCLHb, HbG, and HAR induce HO-1 only HAR-pretreatment improves liver microcirculation. The induction of HO-1 expression and the lack of NO-scavenging through HAR may play an important role in preventing shock-induced sinusoidal perfusion failure.

Effects of S-Nitrosation on Hemoglobin-Induced Microvascular Damage

Blood substitutes, such as diaspirin cross-linked hemoglobin (Hb), cause microvascular leakiness to macromolecules. Because of the potentially stabilizing effects of nitric acid (NO) on endothelium, experiments were performed to determine whether S-nitrosohemoglobin (SNO-Hb), a potential NO-donor Hb-based blood substitute, would not cause microvascular damage. Release of NO, or its metabolites, from the SNO-Hb was facilitated by addition of glutathione, which aids in the decomposition of S-nitrosothiols. In anesthetized rats, the mesenteric microvasculature was perfused with SNO-Hb with glutathione (six rats), SNO-Hb alone (six rats), or saline (eight rats) for 10 min, followed by fluorescein isothiocyanate (FITC)-albumin for 1 min, and finally fixed for epifluorescence microscopic examination. When comparing the SNO-Hb group with saline, both the numbers and areas of leaks were significantly increased [0.019 +/- 0.003 (SEM) microm vs. 0.0030 +/- 0.0004 and 7.36 +/- 1.50 vs. 0.156 +/- 0.035 (p < 0.005)]. With the addition of glutathione, leakage was still high (0.005 +/- 0.00005 microm and 5.086 +/- 0.064 microm) but decreased compared with SNO-Hb alone (p < 0.005). In conclusion, NO, or a related vasodilator, when released from SNO-Hb, significantly reduces but does not eliminate microvascular damage. Further improvements may result by S-nitrosating a more stable form of modified hemoglobin.

Chemical Characterization of Diaspirin Cross-Linked Hemoglobin Polymerized with Poly(ethylene glycol)

A lack of specificity associated with chemical modification methods used in the preparation of certain hemoglobin (Hb)-based oxygen carriers (HBOCs) may alter Hb structure and function, as amino acids located in critical regions (e.g., alpha-beta interfaces and the 2,3-DPG binding pocket) may unintentionally be targeted. Hb protein surface modifications with various poly(ethylene glycol) (PEG) derivatives have been used as conjugating and polymerizing agents with the intent of improving reaction site specificity/reproducibility and ultimately reducing the untoward hypertensive response due to nitric oxide scavenging by smaller molecular size tetrameric species (i.e., 64 kDa) in HBOC solutions. Previous experiments performed in our laboratory have evaluated the influence of polymerization of diaspirin alpha-alpha cross-linked Hb (alphaalpha-DBBF-Hb) with a bifunctional modified PEG, bis(maleoylglycylamide) PEG (BMAA-PEG), in terms of oxygen carrying capacity, redox properties, hypertensive response, and renal clearance in rats. The data presented in this paper specifically evaluate the influence of BMAA-PEG on alphaalpha-DBBF-Hb (Poly-alphaalpha-DBBF-Hb) to identify molecular weight distribution, protein conformation, and site-specific modification, as well as to provide insight into the previously determined in vitro and in vivo functional and vasoactive characteristics of this HBOC. Chemical analysis performed herein reveals nonspecific modifications induced by BMAA-PEG that result in the full modification of alphaalpha-DBBF-Hb leaving no tetrameric cross-linked starting material in solution. These data are inconsistent with the continuing assumption that molecular size (i.e., 64 kDa) has a direct influence on HBOC-mediated vasoactivity and that other protective strategies should be considered to control blood pressure imbalances.

Effects of vitamin C infusion and vitamin E-coated membrane on hemodialysis-induced oxidative stress

Chronic hemodialysis (HD) patients manifest anemia and atherosclerosis with associated oxidative stress. We explored whether intravenous infusion of vitamin C (VC) and/or use of vitamin E (VE)-coated dialysis membrane could palliate HD-evoked oxidative stress. Eighty patients undergoing chronic HD were enrolled and randomly assigned into four groups: HD with intravenous VC (n=20), HD with VE-coated dialyzer (n=20), HD with both (n=20), and HD with neither (n=20). We evaluated oxidative stress in blood and plasma, erythrocyte methemoglobin/ferricyanide reductase (red blood cells (RBC)-MFR) activity, plasma methemoglobin, and pro-inflammatory cytokines in these patients. All patients showed marked increases (14-fold) in blood reactive oxygen species (ROS) after HD. The types of ROS were mostly hydrogen peroxide, and in lesser amounts, O2*- and HOCl. HD resulted in decreased plasma VC, total antioxidant status, and RBC-MFR activity and increased plasma and erythrocyte levels of phosphatidylcholine hydroperoxide (PCOOH) and methemoglobin. Intravenous VC significantly palliated HD-induced oxidative stress, plasma and RBC levels of PCOOH, and plasma methemoglobin levels and preserved RBC-MFR activity. The VE-coated dialyzer effectively prevented RBCs from oxidative stress, although it showed a partial effect on the reduction of total ROS activity in whole blood. In conclusion, intravenous VC plus a VE-coated dialyzer is effective in palliating HD-evoked oxidative stress, as indicated by hemolysis and lipid peroxidation, and by overexpression of proinflammation cytokines in HD patients. Using VE-coated dialyzer per se is, however, effective in reducing lipid peroxidation and oxidative damage to RBCs.

Effects of Diaspirin Crosslinked Hemoglobin (DCLHb) on Microcirculation and Local TissuePO2of Striated Skin Muscle Following Resuscitation from Hemorrhagic Shock

The hemoglobin based oxygen carrier (HBOC) Diaspirin Crosslinked Hemoglobin (DCLHb) has been developed to substitute not only the blood volume, but also to restore the oxygen-carrying properties of blood during hemorrhagic shock. However, it has been suggested that HBOCs may enhance the formation of free oxygen radicals through the release of free iron ions via the Haber-Weiss reaction. The aim of this study was to investigate the effects of DCLHb on the microcirculation, leukocyte-endothelial cell interaction and local tissue oxygenation in striated skin muscle of Syrian golden hamsters during and after resuscitation from hemorrhagic shock. In particular we focused on the local tissue oxygenation after resuscitation with DCLHb (hemoglobin content 10 g%) compared to resuscitation using autologous blood diluted to a hemoglobin content of 10 g%. Hemorrhagic shock was induced for 45 minutes by bleeding the animals at a rate of 33 ml/kg BW maintaining a mean arterial pressure of 30 +/- 5 mmHg. Animals were resuscitated either with 33 ml/kg BW 6% Dextran-60.000 or with 10 g% DCLHb. The control group received shed blood diluted with Ringers to a hemoglobin content of 10 g%. Intravital microscopy was used for investigation of the microcirculatory parameters and a multiwire platinum surface electrode for measurement of local tissue pO2 in striated skin muscle in the dorsal skinfold chamber of Syrian golden hamsters. Resuscitation from hemorrhagic shock with 10 g% AUB revealed significant increase of leukocytes rolling in postcapillary venules at 30 to 120 minutes after resuscitation compared to baseline values. DCLHb turned out to reduce the number of firmly adherent leukocytes after resuscitation compared to 10 g% AUB. Microvascular permeability as an indicator for functional endothelial integrity revealed no significant differences between the groups. DCLHb and 10 g% AUB led to a significant increase in local tissue oxygenation after resuscitation from hemorrhagic shock. However, 10 g% AUB turned out to be most effective to restore the local tissue pO2 compared to Dx-60. Our findings indicate that DCLHb restores microvascular perfusion after critical hemorrhagic shock as efficient as Dx-60 and 10 g% AUB. The absence of enhanced leukocyte-endothelium interaction after resuscitation with DCLHb implies that this HBOC does not exacerbate formation of oxygen free radicals during reperfusion. DCLHb effectively increases local tissue pO2 after resuscitation from hemorrhagic shock; however, not as effectively as 10 g% AUB.