Effect of polymyxin B on hemoglobin-mediated hepatotoxicity.

In previous studies, enhanced toxicity of hemoglobin when combined with endotoxin has suggested a specific binding site. This binding may take place even when endotoxin-free hemoglobin is administered, due to some low levels of endotoxin usually present in the portal vein. To test this hypothesis, we administered endotoxin-free, stroma-free human hemoglobin (SFH) to rabbits in doses shown previously to be hepatotoxic or lethal. A second group of rabbits received the anti-endotoxin antibiotic polymyxin B (PB) to inactivate endotoxin, followed by a dose of SFH. A control group received intravenous PB alone. There was no differences in the incidence of hepatotoxicity among these three groups. To show the binding interactions of endotoxin with PB and endotoxin with hemoglobin, an experiment using toluidine blue was performed. According to this study, specific binding of hemoglobin with endotoxin did not occur. Our results do not agree with previous suggestions that hemoglobin-mediated hepatotoxicity is due to the interaction in vivo of hemoglobin with low levels of endogenous endotoxin.

Effect of pyridoxalated stabilized stroma-free hemoglobin solution on the clearance of intravascular lipid by the reticuloendothelial system.

Although considerable work is being carried out with pyridoxalated glutaraldehyde-stabilized stroma-free hemoglobin solution (SFHS-pp) as a possible blood substitute, it is not known whether the SFHS-pp has any adverse effect on some aspects of reticuloendothelial (RE) function. To determine this, 131I-gelatinized trioleate (GT) was injected 30 min or 3 days after the injection of 45 or 100 mg/100 gm BW SFHS-pp (low and high dose, respectively). The results indicate that low as well as high dose SFHS-pp significantly decreased splenic uptake and transiently increased pulmonary uptake of GT. Hepatic retention of GT, however, increased 3 days following SFHS-pp administration, indicating that there was a stimulation of this aspect of liver RE function at that time. The intravascular clearance rates of GT at 3 days were also markedly faster, but no dose effect relationship was seen. This suggests that there was an overall stimulation of RE function 3 days following SFHS-pp administration. These results provide support for the safety of polymerized stroma-free hemoglobin solution for use as an oxygen-carrying volume support.

Pelvic hematoma after intercourse while on chronic anticoagulation.

A 41-year-old anticoagulated woman presented with increasing pelvic pain that had begun two days prior during intercourse. An ultrasound and computerized tomographic (CT) scan confirmed the diagnosis of pelvic hematoma. The hematoma resolved spontaneously with normalization of clotting studies. The emergency physician should consider this rare cause of pelvic pain in selected anticoagulated patients. The need for a thorough history in emergency patients is emphasized, as well as the value of ultrasound and CT scan in the diagnosis of this disorder.

Deleterious effects of stroma-free hemoglobin used as resuscitative fluid for rats with ischemic intestinal shock.

This study examined the effect of modified stroma-free Hgb solution (SFHS) as a resuscitative fluid for rats in an ischemic intestinal shock model. The experiments compared the effect of SFHS with two other resuscitative fluids: lactated Ringer's solution (RL) and 3% albumin in RL. The control group received no fluids. After the induction of shock, the resuscitative solution was given iv over a 6-h period. RL and albumin in RL resulted in long-term (greater than 7 days) survival in 15 of 16 animals. All animals in the control group and six of eight animals in the SFHS-treated group died within 24 h. Hct was used to estimate changes in intravascular volume. All control animals became hemoconcentrated and exhibited a 50% loss in plasma volume. The failure of SFHS to improve significantly the survival in rats compared to the controls is surprising in view of the fact that the SFHS had the same oncotic pressure as the albumin-containing solution and identical volumes were administered.

Removal of bacteria from blood by charcoal hemoperfusion.

E. coli bacteria were successfully removed from contaminated RBC/plasma by using a special matrix of micro-encapsulated albumin activated charcoal (ACAC). Efficacy of removing the bacteria was directly related to the amount of time the contaminated blood was in contact with the charcoal. The data indicated that the bacteria adhered to the ACAC, but that the charcoal was not bactericidal.

Graft rejection in earthworms: an electron microscopic study.

Graft rejection in the earthworm, Lumbricus terrestris, is mediated by granulocytic coelomocytes (neutrophils, by light microscopy). Within 1--3 days post-transplantation of body-wall xenografts, granulocytic coelomocytes migrate to the transplant site, penetrate the graft matrix, and actively phagocytize viable muscle fibers. Destruction of muscle tissues may be augmented by lymphocytic coelomocytes (basophils, by light microscopy), but these cells participate in a minor way and do so very late in the rejection process (days 11--13). Xenografts, autografts and sham-operated controls display generalized inflammatory reactions, including coelomyocyte infiltration into the grafted tissues. In autografts, however, granulocytes only penetrate the inner longitudinal muscle layers and extensive infiltration, persistence and complete destruction never occur. Lymphocytic coelomocytes are not observed in autografts or sham-operated controls. Grafts rejection by leukocytes in an advanced invertebrate confirms that aspects of cell-mediated immunity evolved early in phylogenetic history.

Electron-microscopic observations of normal coelomocytes from the earthworm, Lumbricus terrestris.

Coelomocytes of the earthworm, Lumbricus terrestris, were studied by transmission electron microscopy. Four morphological cell types are distinguishable: lymphocytic coelomocytes, granulocytic coelomocytes, eleocytes (chloragogen cells), and inclusion-containing coelomocytes. Within these major categories, several distinct cell types differ and may represent developmental stages. The two types of lymphocytic coelomocytes are small with central nuclei and scanty cytoplasms. Two types of granulocytic coelomocytes differ greatly in shape and content; both have small dark-staining granules that resemble lysosomes. Electrocytes, derived from chloragogen tissue, contain a variety of granules, inclusions and vacuoles. Inclusion-containing coelomocytes appear as two types which may be immature and mature forms. Although these cells resemble those that have been referred to as erythroid cells in other invertebrates, the large inclusion bodies are apparently unrelated to hemoglobin; they can undergo morphologic transformation and be extruded by exocytosis. This information on lymphocytic, granulocytic and inclusion-containing coelomocytes is crucial to understanding more about cellular immunity in the earthworm.

Earthworm coelomocytes in vitro.

Contamination and low viability of earthworm coelemocytes in tissue culture have delayed in vitro studies. Using penicillin, streptomycin, tetracycline and Amphotericin B, Lumbricus terrestis coelomocytes were maintained viable and uncontaminated for 10 days at 15degreesC in medium L-15 supplemented with 5 to 10% fetal bovine serum. The coelomocytes survived for at least 10 days with 85% viability as assessed by trypan blue exclusion assays and phagocytosis of heat-killed yeast. Studies on the thymidine uptake, however, were negative. With the involvement of coelomocytes in tissue graft rejection, in vitro techniques can now be applied to study their capacity in the immune response.

Safety and immunogenicity of a recombinant outer surface protein A Lyme vaccine.

OBJECTIVE: To evaluate the safety and immunogenicity of a recombinant outer surface lipoprotein A (OspA) Lyme vaccine in healthy adults. DESIGN: Randomized, double-blind, placebo-controlled trial. SETTING: Clinical research unit of a medical center. PARTICIPANTS: Thirty-six healthy adult volunteers aged 18 through 65 years. INTERVENTIONS: Volunteers were randomly assigned to receive two 10-micrograms doses of OspA Lyme vaccine, OspA Lyme vaccine adsorbed to alum, or a buffer placebo. Subjects in the OspA Lyme vaccine group received a third dose. Patients were assessed after each vaccination for a total follow-up period of 1 year. Serum samples for antibody determination were drawn at baseline, 2 and 3 weeks after dose 1, once per week for 4 weeks after dose 2, 20 weeks after dose 2, and 1 month after dose 3. MAIN OUTCOME MEASURES: Local and systemic adverse reactions and antibody levels specific for OspA. RESULTS: The most common reactions were local pain and tenderness at the injection site. Adverse events did not increase following the second or third dose. Two doses of both vaccine formulations elicited high-titer antibodies that inhibited replication of Borrelia burgdorferi in vitro. No differences were noted in antibody levels elicited by the adsorbed and nonadsorbed formulations. CONCLUSION: Two or three doses of OspA Lyme vaccine are safe and immunogenic in adults.

Ascorbate-2-phosphate in red cell preservation. Clinical trials and active components.

A red cell additive solution (AS-005) containing ascorbate-2-phosphate (AsP) to maintain 2,3-diphosphoglycerate, plus adenine, phosphate, and mannitol to retain viability and reduce hemolysis, was evaluated by human clinical trials. A crossover design was used with another additive solution (Nutricel AS-3, Cutter Laboratories) serving as the control for each donor. Each additive solution was evaluated at 35 and 42 days of storage. There was no significant difference between the red cell viability of the two storage solutions at either time period. Split-bag, AS-005 in vitro studies at two temperatures (2.5 and 5.5 degrees C), both within the range of 1 to 6 degrees C approved by the American Association of Blood Banks and the Food and Drug Administration, resulted in dramatically different in vitro parameters, including a threefold difference in 2,3-diphosphoglycerate (2,3-DPG), a fivefold difference in glucose, and significant differences in pH and adenosine triphosphate. High-pressure liquid chromatography data confirmed the preliminary report that 1 to 2 percent (wt/wt) oxalate was present in preparations of AsP. In vitro storage data confirmed that oxalate is the active component of AsP that preserves 2,3-DPG during storage.

Stroma-free methemoglobin solution as an antidote for cyanide poisoning: a preliminary study.

Effective treatment of cyanide poisoning requires rapid diagnosis, good supportive treatment and the use of a specific antidote. The currently available antidotes offer demonstrated efficacy along with significant potential adverse side effects. We have investigated an alternate approach to antidote therapy for cyanide poisoning by using Stroma-Free Methemoglobin Solution ( SFMS ). Rats injected with an LD100 intravenous dose of cyanide were treated with SFMS equal to 1.5% of their total body hemoglobin. There was a highly significant increase in the survival rate of the treated group compared to saline controls. The potential advantages of SFMS over current antidotes include an immediate onset of action, rapid elimination of cyanide from the body and a mode of action that doesn't compromise any of the patients' oxygen carrying capacity. SFMS shows promise as a significant adjunct in the treatment of cyanide poisoning.

Safety and immunogenicity of NYVAC-JEV and ALVAC-JEV attenuated recombinant Japanese encephalitis virus--poxvirus vaccines in vaccinia-nonimmune and vaccinia-immune humans.

A controlled, randomized, double-blind clinical trial evaluated whether two attenuated recombinant poxviruses with identical Japanese encephalitis virus (JEV) gene insertions, NYVAC-JEV and ALVAC-JEV, were safe and immunogenic in volunteers. Groups of 10 volunteers distinguished by vaccinia immune status received two doses of each vaccine. The vaccines appeared to be equally safe and well tolerated in volunteers, but more reactogenic than licensed formalin-inactivated JE and placebo vaccines given as controls. NYVAC-JEV and ALVAC-JEV vaccine recipients had frequent occurrence of local warmth, erythema, tenderness, and/or arm pain after vaccination. There was no apparent effect of vaccinia immune status on frequency or magnitude of local and systemic reactions. NYVAC-JEV elicited antibody responses to JEV antigens in recipients but ALVAC-JEV vaccine poorly induced antibody responses. However, NYVAC-JEV vaccine induced neutralizing antibody responses only in vaccinia-nonimmune recipients while vaccinia-immune volunteers failed to develop protective antibodies (5/5 vs. 0/5 seroconversion, p<0.01). These data suggest that preexisting immunity to poxvirus vector may suppress antibody responses to recombinant gene products.