Diaspirin Cross-linked Hemoglobin Solution Research Articles

EFFECT OF DIASPIRIN CROSSLINKED HEMOGLOBIN (DCLHb HemAssist™) DURING HIGH BLOOD LOSS SURGERY ON SELECTED INDICES OF ORGAN FUNCTION*

The safety of the hemoglobin based oxygen carrier diaspirin crosslinked hemoglobin (DCLHb) has been reported only in the low (50-200 mg/kg) dose range [Przybelski. R.J.; Daily, E.K.; Kisicki, J.C.; Mattia-Goldberg, C.; Bounds, M.J.; Colburn, W.A. Phase I study of the safety and pharmacologic effects of diaspirin crosslinked hemoglobin solution. Crit. Care Med. 1996, 24 (12), 1993-2000, Bloomfield, E.; Rady, M.; Popovich, M.; Esfandiari, S.; Bedocs, N. The use of diaspirin crosslinked hemoglobin (DCLHb 1996, 95, (3A), A220.]. We conducted a randomized prospective open-label trial of DCLHb and packed red blood cells (PRBCs) in high-blood loss surgical patients to show the effect of 750 ml DCLHb (approximately 1000 mg/kg) on selected indices of organ function. After institutional approval, 24 patients scheduled to undergo elective orthopedic or abdominal surgery, were randomized to receive either PRBCs or 10% DCLHb within 12 hours after the start of surgery. Patients with renal insufficiency, abnormal liver function, severe coronary artery disease (CAD) and ASA physical status > or = IV were excluded. The anesthetic technique was left to the judgment of the anesthesiologist. Autologous predonation and intraoperative blood conservation techniques were utilized as appropriate. The indications for blood transfusion were individualized on disease state, stage of surgery, and plasma Hb concentration. Laboratory studies were obtained preoperatively and up to 28 days postoperatively. Patients were observed daily for development of jaundice, hematuria, nausea, vomiting, gastrointestinal discomfort, cardiac, respiratory, and infectious complications. Organ effects were assessed with urinalysis, creatinine clearance, electrocardiogram (ECG), and a panel of blood and serum laboratory tests. The dose of DCLHb administered ranged from 680-1500 mg/kg (mean = 999 mg/kg). Estimated blood loss was 27 +/- 13 ml/kg and 31 +/- 15 ml/kg in the control and DCLHb groups, respectively. Fewer PRBCs (1.9 +/- 1.2 vs. 3.4 +/- 2.4 units. P = 0.06) were transfused to DCLHb patients on the operative day although this difference was no longer apparent later on. In the DCLHb group, 4/12 patients avoided any allogeneic PRBC transfusion vs. none in the control group (P = 0.09). Systolic, diastolic and mean blood pressure increased moderately after DCLHb for a period of 24-30 hours. There were no occurrences of cardiac ischemia. myocardial infarction, stroke, or pulmonary edema, by clinical or laboratory parameters up to the 28th postoperative day (POD). Seven of 12 (58%) DCLHb patients had yellow skin discoloration vs. none in the PRBC group (P < 0.01). Two of four non-urologic surgery patients developed asymptomatic postoperative hemoglobinuria after DCLHb. Creatinine clearance was unchanged postoperatively. Because of hemoglobin interference, bilirubin, gamma-glutamyl transferase (GGT), and amylase could not be measured reliably on POD1; on POD2. amylase was transiently elevated to 3 times ULN along with mild elevations of bilirubin, transaminases and BUN. Mean total creatine phoshokinase (CPK) peaked at 8 times the upper limit of normal (ULN) in the DCLHb group, compared with less than twice ULN for controls. Three DCLHb patients had prolonged ileus. Two of these patients had postoperative hyperamylasemia, one of whom developed mild pancreatitis. DCLHb did not affect white blood cell count or coagulation tests. Administration of approximately 1000 mg/kg DCLHb was associated with transient arterial hypertension, gastrointestinal side effects, laboratory abnormalities, yellow skin discoloration, and hemoglobinuria. These observations point to opportunities for improvement in future synthetic hemoglobin design.

Randomized trial of diaspirin cross-linked hemoglobin solution as an alternative to blood transfusion after cardiac surgery: M.L. Lamy, E.K. Daily, J.F. Brichant, et al, for the DCLHb Cardiac Surgery Trial Collaborative Group. Anesthesiology 92:646–656, 2000

Randomized trial of diaspirin cross-linked hemoglobin solution as an alternative to blood transfusion after cardiac surgery: M.L. Lamy, E.K. Daily, J.F. Brichant, et al, for the DCLHb Cardiac Surgery Trial Collaborative Group. Anesthesiology 92:646–656, 2000

Diaspirin-crosslinked hemoglobin reduces mortality of severe hemorrhagic shock in pigs with critical coronary stenosis

To evaluate the effects of resuscitation with a 10% diaspirin-crosslinked hemoglobin (DCLHb) solution on global hemodynamic variables, systemic and myocardial oxygen transport and tissue oxygenation, and contractile function of the left ventricle in an experimental model of severe hemorrhagic shock and critical stenosis of the left anterior descending coronary artery (LAD). Prospective, placebo-controlled, randomized study. Experimental animal laboratory. A total of 20 anesthetized pigs. After implementation of a permanent critical LAD stenosis (ie, maintenance of basal blood flow but absence of reactive hyperemia after a 10-sec complete vessel occlusion), hemorrhagic shock (target mean aortic pressure, 45 mm Hg) was induced within 15 mins by programmed withdrawal of blood and maintained for 60 mins. Subsequently, the volume of plasma lost during hemorrhage was replaced by either a balanced electrolyte solution containing 10 g/dL DCLHb (DCLHb group; n = 10) or an 8 g/dL human albumin solution (HSA) oncotically matched to DCLHb (HSA group; n = 10). Data were collected immediately after the infusion of the different solutions and again after 60 mins had elapsed. Although five of ten HSA-treated animals died of acute left ventricular failure within the first 20 mins after complete fluid resuscitation, all of the DCLHb-treated animals survived the 60-min observation period after resuscitation (p < .05). This significant difference in mortality is explained by higher coronary perfusion pressure in DCLHb-treated animals (75 +/- 17 vs. 27 +/- 17 torr DCLHb vs. HSA group; p < .05) and persistence of subendocardial ischemia and hypoxia (radioactive microspheres method) in HSA-treated animals on resuscitation particularly affecting the LAD-supported myocardium (subendocardial oxygen delivery: 20 +/- 11 vs. 3 +/- 1 mL oxygen x g(-1) x min(-1), DCLHb vs. HSA group; p < .05). Except for enhanced myocardial contractility immediately on infusion of DCLHb (maximal left ventricular pressure increase: 2373 +/- 782 vs. 1730 +/- 543 torr x sec(-1) DCLHb vs. HSA group; p < .05), no differences were detected between groups concerning the variables of systemic oxygen transport, tissue oxygenation, and regional contractile function of the myocardium (determined with microsonometry). Fluid resuscitation with 10% DCLHb solution completely reverses hemorrhagic shock-induced subendocardial ischemia and hypoxia in the presence of compromised coronary circulation and thereby prevents early death after resuscitation.

Diaspirin crosslinked hemoglobin (DCLHb) attenuates bacterial translocation in rats.

Intestinal barrier function is compromised following severe hemorrhage which may allow bacterial translocation (BT) to occur and subsequently initiate a systemic response leading to multiple system organ failure (MSOF). This study compared BT following hemorrhage and resuscitation with lactated Ringer's solution (LR) or diaspirin crosslinked hemoglobin solution (DCLHb). Rats (250-350 grams) were hemorrhaged to a base deficit of 15 +/- 2 mmol/L and immediately resuscitated with either 3:1 LR or 1:1 DCLHb based on shed blood volume. Four hours following resuscitation, the mesenteric lymph node complex was harvested, homogenized and plated onto MacConkey and Columbia CNA agar culture media. Facultative anaerobic and obligate aerobic bacteria were identified 48 hours later in 11/22 (50%) LR-treated rats and in 4/21 (19%) DCLHb-treated rats (p < or = 0.05). Following resuscitation, base excess (BE) and central venous oxygen saturation (SvO2) were not only restored to baseline but were significantly greater (p < or = 0.05) in DCLHb-treated rats than in LR-treated rats. In a separate group of rats subjected to the same hemorrhage and resuscitation protocol, mean arterial pressure in DCLHb-treated rats, but not LR-treated rats, was restored to baseline by 15 minutes and remained at or above baseline for up to 4 hrs. Twenty-four hour survival was 50% in LR-treated rats and 77% in DCLHb-treated rats (p > 0.05). These data suggest that DCLHb is superior to LR in restoring tissue oxygen delivery, as judged by BE and SvO2. Furthermore, since DCLHb restores oxygen delivery and attenuates BT, early resuscitation with DCLHb may limit gut ischemia and subsequent gut barrier failure and hence prevent the development of sepsis, MSOF and subsequent death.

THE EFFICACY OF DIASPIRIN CROSSLINKED HEMOGLOBIN SOLUTION RESUSCITATION IN A MODEL OF UNCONTROLLED HEMORRHAGE

Controversy exists whether early aggressive fluid therapy in the setting of uncontrolled hemorrhage worsens outcome by increasing blood loss from injured vessels. Since diaspirin crosslinked hemoglobin (DCLHb) is a vasoactive, oxygen-carrying solution, we compared the effects of DCLHb with other resuscitative fluids on blood loss, hemodynamics, and tissue oxygen delivery in a model of uncontrolled hemorrhage. Anesthetized rats (250-350 g) were subjected to a 50% tail transection and resuscitated 15 minutes later with 1:1 DCLHb, 3:1 lactated Ringer's solution (LR), 1:1 hypertonic saline (7.5% HTS), or 1:1 human serum albumin (8.3% HSA) based on initial volume of blood loss (average 4.7 +/- 0.3 mL/kg). An unresuscitated group served as a control. Cumulative blood loss was measured at 5 hours postresuscitation. By 15 minutes after tail transection, mean arterial pressure (MAP) decreased 19.2 +/- 3.8 mm Hg from the baseline value (102 +/- 5 mm Hg). The DCLHb solution restored and maintained MAP and subcutaneous tissue oxygen tension at baseline values better than all other resuscitative fluids. Although blood loss in DCLHb-treated animals was greater than in unresuscitated animals, it was no different from other resuscitative fluids and less than with HSA. There was no difference in 24-hour survival between all treatment groups. In conclusion, DCLHb elevates MAP but does not exacerbate blood loss or compromise tissue oxygen delivery compared with other resuscitative fluids in this model of uncontrolled hemorrhage.

Characterization of the hemodynamic response to intravenous diaspirin crosslinked hemoglobin solution in rats.

Diaspirin crosslinked hemoglobin solution (DCLHB) has potential for clinical use as an oxygen-carrying solution because of its excellent oxygen transport properties and biochemical stability. The present study characterizes the effects of intravenous infusions of 0.625-40 mL/kg (62.5-4000 mg/kg) DCLHb on mean blood pressure (MAP) and heart rate (HR) in conscious rats. DCLHb at all doses tested except 62.5 mg/kg was associated with an immediate increase in MAP (25-30% above baseline) that peaked between 20-30 minutes after infusion and returned to baseline within 120-300 minutes in a dose-dependent manner. Maximum MAP achieved was in the range of 129 +/- 7 to 140 +/- 7 mm Hg and there was no statistically significant difference in the response between doses. HR responded in a reciprocal manner to changes in MAP. Volume- and oncotic-matched infusions of LR and albumin did not alter MAP or HR. Slow infusion (0.34 mL/min) of DCLHb appeared to blunt the magnitude of the pressor response when compared to bolus injection (< 10 sec). DCLHb administration is associated with a pressor response that is not due to volume load, oncotic pressure, or rate of infusion, suggesting that it is intrinsic to the modified hemoglobin molecule and pharmacologic in nature.

Diaspirin Cross-Linked Hemoglobin Solution as a Resuscitatwe Fluid Following Severe Hemorrhage in the Rat

The effect of diaspirin cross-linked hemoglobin (DCLHb) on mean arterial pressure (MAP) and heart rate (HR) was compared to Ringer's lactate (RL) and shed blood in a 70% lethal model of hemorrhage (35 cc/kg blood loss) in conscious rats. All animals resuscitated with DCLHb regardless of dose (17.5 and 35 cc/kg) and concentration (7% and 10% solution) exhibited complete restoration of MAP and HR which was maintained for at least 5 hrs. Hemodynamic responses in DCLHb-treated animals were not significantly different from 35 cc/kg blood-treated animals. In RL (105 cc/kg) treated animals the MAP was restored to 60-70% of baseline. 24 hr survival in animals resuscitated with fluids ranged between 88-100% and was not significantly different between treatment groups.

Immunogenicity of diaspirin cross-linked human hemoglobin solutions.

To assess the potential immunogenicity of human diaspirin cross-linked hemoglobin (DCLHb) solution, repetitive doses of this material were given intravenously to rhesus monkeys at monthly intervals and the immune response to this challenge was assessed. Serum samples collected at multiple intervals throughout the study showed no evidence of DCLHb specific IgG or IgM production. Intradermal skin tests performed one month after the final DCLHb infusion were also negative. These data demonstrate that DCLHb is not antigenic when administered intravenously to rhesus monkeys. In addition, screening of a panel of normal human sera for pre-existing anti-DCLHb IgG antibodies was negative, suggesting that this modified hemoglobin is unlikely to be antigenic in humans.

Rat maze performance after infusion of cross-linked hemoglobin solution.

Neurotoxicity and behavioral performance degradation have previously been observed in rats after exchange transfusions with unmodified stroma-free hemoglobin solutions. We evaluated a diaspirin cross-linked stroma-free hemoglobin solution (HbXL) for evidence of neurotoxicity. Rats that were trained to complete a water alley maze received a clinically-relevant dose (20 ml/kg) of the 14% HbXL solution on top of their normal blood volume. After post-treatment memory testing in the water maze, the same rats were challenged to learn an elevated radial-arm maze. The HbXL and control groups showed no water maze performance degradation after treatment, and all groups demonstrated learning of the radial-arm maze as shown by decreased errors and times to completion. The brains, heart, and livers presented normal histology thirty days after infusion, but six of fifteen animals showed marked renal tubular regeneration. The normal memory and learning performance and brain histology after infusion with HbXL suggests that this hemoglobin solution is not neurotoxic to unhemorrhaged rats.