Pharmacologic interactions of exogenous lung surfactant and recombinant human Cu/Zn superoxide dismutase.

The effect of exogenous surfactant on the pharmacokinetics of intratracheally administered recombinant human superoxide dismutase (rhSOD) was studied. Five groups of rats received the following intratracheally: 1 mL/kg of saline; 5 or 25 mg/kg of rhSOD; or 4 mL/kg of exogenous surfactant followed in 30 min by 5 or 25 mg/kg of rhSOD. Animals were killed at 24, 48, and 72 h, and serum, bronchoalveolar lavage, and lung tissue were analyzed for rhSOD. rhSOD was not detected in the lungs of saline-treated animals or in serum from any animals. At 24 h, lung-tissue rhSOD was higher in rats treated with surfactant and rhSOD versus rhSOD alone (5 mg/kg: 6.8 +/- 2.5 versus 0 microgram/whole lung, p < 0.05; 25 mg/kg: 29.9 +/- 9.6 versus 0.1 +/- 0.1 microgram/whole lung, p < 0.05). Bronchoalveolar lavage fluid levels correlated well with lung tissue concentrations. By 48 h, lung tissue rhSOD concentrations were insignificant in all groups. rhSOD was still present in lavage fluid from rats treated with surfactant and rhSOD. No rhSOD was detected at 72 h. In separate in vitro experiments, physical and biological drug-drug interaction studies were performed. When radiolabeled rhSOD was combined with exogenous surfactant and centrifuged at 10,000 x g for 30 min, 81.3 +/- 2.5% of rhSOD was found in the supernatant versus 18.7 +/- 2.5% in the surfactant pellet. Serial washing of the surfactant pellet removed virtually all remaining rhSOD. This finding suggests that the rhSOD and surfactant were only weakly associated. Combining rhSOD and exogenous surfactant did not alter the activity of either agent.(ABSTRACT TRUNCATED AT 250 WORDS)

Active oxygen species play a role in mediating platelet aggregation and cyclic flow variations in severely stenosed and endothelium-injured coronary arteries.

A canine model with cyclic flow variations (CFVs) in stenosed and endothelium-injured coronary arteries was used to examine the role of active oxygen species in platelet aggregation in vivo. We studied 90 anesthetized dogs in which the pericardial cavity was opened and the heart was exposed. The velocity of blood flow in the left anterior descending coronary artery (LAD) was monitored by a pulsed Doppler flow probe. In 67 dogs, the LADs were stenosed by applying external constrictors at the site where the endothelium was mechanically injured. CFVs developed in all 67 dogs. Treatment with the antioxidants recombinant human copper-zinc superoxide dismutase (r-h-CuZnSOD), recombinant human manganese superoxide dismutase (r-h-MnSOD), and catalase eliminated platelet aggregation-associated coronary CFVs in 63%, 62%, and 64% of animals, respectively. Intravenous infusion of epinephrine restored CFVs in most dogs. Ketanserin, a serotonin (5-hydroxytryptamine2) receptor antagonist, abolished epinephrine-restored CFVs and eliminated CFVs in dogs in which CFVs had not been eliminated by free radical scavengers. In an additional 23 dogs, the LADs were stenosed but not mechanically injured. For control studies, saline was infused into the LADs of 5 dogs. Xanthine/xanthine oxidase was infused into the LADs of 8 dogs and induced CFVs in 4. Hydrogen peroxide was infused into the other 10 dogs and induced CFVs in 9. Histological analysis of the coronary artery revealed that the intima was significantly injured by the infusion. In ex vivo platelet aggregation studies, the in vivo treatment with r-h-CuZnSOD, r-h-MnSOD, and catalase significantly inhibited platelet aggregation induced by platelet-activating factor. Thus, active oxygen species are involved in mediating platelet aggregation and cyclic flow variations in stenosed and endothelium-injured canine coronary arteries in vivo.

Prophylactic effects of recombinant human superoxide dismutase in neonatal lung injury.

To determine if recombinant human Cu-Zn superoxide dismutase (rhSOD) would prevent acute lung injury caused by hyperoxia and barotrauma, 26 newborn piglets were studied. Ten piglets were hyperventilated (arterial PCO2 15-20 Torr) with 100% O2 for 48 h. A second group received identical treatment for 4 h (n = 2) or 48 h (n = 8) but was given 5 mg/kg of rhSOD intratracheally at time 0. Six piglets were normally ventilated (arterial PCO2 40-45 Torr) for 48 h with 21% O2. Pulmonary function and tracheal aspirates were examined at time 0 and at 24 and 48 h, and bronchoalveolar lavage was performed at 48 h. In piglets treated with hyperoxia and hyperventilation, lung compliance decreased 42%, and tracheal aspirates showed an increase in neutrophil chemotactic activity (32%), total cell counts (135%), elastase activity (93%), and albumin concentration (339%) over 48 h (P < 0.05). All variables were significantly lower in rhSOD-treated piglets and comparable to normoxic control values. Surfactant remained active in all groups. Immunohistochemistry demonstrated that at 48 h significant rhSOD was distributed homogeneously in terminal airways. Adding rhSOD to tracheal aspirates of hyperoxic hyperventilated piglets did not alter neutrophil chemotaxis, suggesting that rhSOD protected the lung by reducing the production of chemotactic mediators. Results indicate that acute lung injury caused by 48 h of hyperoxia and hyperventilation is significantly ameliorated by prophylactic intratracheal administration of rhSOD.

Growth hormone and insulin reverse net whole body and skeletal muscle protein catabolism in cancer patients.

The authors examined the effect of recombinant-human growth hormone (r-hGH) and insulin (INS) administration on protein kinetics in cancer patients. Twenty-eight cancer patients either received r-hGH for 3 days (GH group, n = 12, weight loss = 6 +/- 2%) or were not treated (control [CTL] group, n = 16, weight loss = 11 +/- 2%) before metabolic study. Recombinant-human growth hormone dose was 0.1 mg/kg/day (n = 6) or 0.2 mg/kg/day (n = 6). Patients then underwent measurement of baseline protein kinetics (GH/B, CTL/B) followed by a 2-hour euglycemic insulin infusion (1 mU/kg/minute) and repeat kinetic measurements (GH/INS,CTL/INS). Whole-body protein net balance (mumol leucine/kg/minute) was higher (p less than 0.05) in GH/INS (0.20 +/- 0.06) than in CTL/INS (0.06 +/- 0.03) or GH/B (-0.19 +/- 0.03). Skeletal muscle protein net balance (nmol phenylalanine/100 g/minute) in GH/INS (25 +/- 6) and CTL/INS (19 +/- 5) was higher than CTL/B (-18 +/- 3). Recombinant-human growth hormone and insulin reduce whole-body and skeletal muscle protein loss in cancer patients. Simultaneous use of these agents during nutritional therapy may benefit the cancer patient.

Growth hormone and insulin combine to improve whole-body and skeletal muscle protein kinetics.

BACKGROUND: A cooperative effect of exogenous insulin and recombinant human growth hormone (r-hGH) with respect to whole-body and skeletal muscle protein metabolism has not been demonstrated previously. This study examined the effect of r-hGH and insulin administration during euglycemic clamping and concurrent amino acid supplementation. METHODS: Twenty-three normal volunteers in the postabsorptive state were either treated with r-hGH for 3 consecutive days before a metabolic study (GH group; n = 10) or not treated (CTRL group; n = 13). The r-hGH dose was 0.2 mg/kg/day (n = 5) or 0.1 mg/kg/day (n = 5). All subjects then received an infusion of 14C-labeled leucine and tritiated phenylalanine, followed by measurement of baseline protein kinetics (GH and CTRL). Subsequently a euglycemic insulin infusion (1 mU/kg/min) with concurrent amino acid infusion was administered, and protein kinetic measurements were repeated at steady state. RESULTS: GH and insulin separately produced an increase in whole-body and skeletal muscle protein net balance. GH plus insulin was associated with a higher net balance of protein than was insulin alone. CONCLUSIONS: r-hGH and insulin in the presence of amino acids and glucose combine to improve whole-body and skeletal muscle protein kinetics.

Effect of recombinant human growth hormone on acute and chronic human immunodeficiency virus infection in vitro.

Growth hormone (somatotropin) is a potent anabolic protein currently being evaluated clinically in cachexia associated with malignancy and human immunodeficiency virus (HIV) disease. Growth hormone can also lead to enhancement of lectin-mediated cellular proliferation, macrophage activation, and cytokine induction, events linked to induction of latent HIV in vitro. We thus explored the ability of recombinant human growth hormone (rhGH) to affect viral replication in acute and chronic HIV infection, and to alter transcription at the HIV-1 long terminal repeat (LTR). A clone of promonocytic cells, chronically infected with HIV-1 and susceptible to viral induction by a variety of cytokines and protein kinase C activators, was unperturbed by rhGH used over broad concentrations (10 to 500 ng/mL) and time intervals. This unresponsiveness paralleled the lack of effect of rhGH on HIV-associated trans-activation in both monocytic and CD4+ T-cell lines. In contrast, rhGH enhanced viral replication in acutely infected peripheral blood mononuclear cells (PBMC) by twofold to 20-fold, albeit having no adverse effect on the antiviral efficacy of zidovudine (AZT). Augmentation of HIV growth correlated with stimulation of cellular DNA synthetic responses and an increase in tumor necrosis factor-alpha (TNF-alpha) secretion. These data are discussed in the context of ongoing clinical trials of rhGH in HIV-seropositive individuals with wasting syndromes.

Pharmacokinetics of recombinant human superoxide dismutase.

Superoxide dismutase (SOD) disposition was studied in order to design a rational approach for drug administration in the setting of acute myocardial infarction. Four chronically instrumented conscious dogs received the following dosage regimens of recombinant human SOD (rhSOD) on successive days: (a) 5 mg/kg left atrial (LA) bolus, (b) 5 mg/kg central vein (CV) bolus, (c) 15 mg/kg CV bolus, and (d) 5 mg/kg CV infusion over 60 min; additionally, all dogs received (e) a 5 mg/kg CV bolus under pentobarbital anesthesia. Serial serum samples were obtained after each dose and serial myocardial samples were obtained after dose (e). The serum rhSOD concentration was measured by radioimmunoassay and the data were fit to a two-compartment model. The distribution half-life was 7.8 +/- 1.7 min (mean +/- SEM), and the elimination half-life was 51.1 +/- 5.9 min; the central compartment volume of distribution (Vc) was 81 +/- 26 ml/kg and the steady-state volume of distribution was 156 +/- 20 ml/kg. The dosage regimen had no influence on clearance rates. Peak plasma concentrations (micrograms/ml) for the dosage regimens were (a) 65 +/- 28, (b) 89 +/- 19, (c) 214 +/- 61, (d) 20 +/- 5, and (e) 86 +/- 9. The peak level following continuous infusion did not occur until 50 min of infusion and was only one-fourth of the level achieved with a bolus of the same dose. Myocardial levels were less than 1% of serum levels, suggesting negligible rhSOD penetration into the myocardium.(ABSTRACT TRUNCATED AT 250 WORDS)

Carbodiimide modification of superhelical PM2 DNA: considerations regarding reaction at unpaired bases and the unwinding of superhelical DNA with chemical probes.

Superhelical PM2 DNA I can be modified with N-cyclohexyl-N'-beta-(4-methylmorpholinium)ethyl carbodiimide (CMC). The transition of the sedimentation coefficient uncorrected for buoyant density change (S20,*) vs. % reactivity in terms of base pairs shows the following characteristics. The S20,* increases by 4.5 S units upon 1% modification. There is a plateau in S20,* between 1 and 4% reactivity. The extent of reactivity was determined by buoyant density and 14C radioactive CMC binding measurements. Further reactivity was not explored since Pulleyblank and Morgan's (22) data of S20,* vs. % reactivity from 6 to 34% was previously published. The initial results obtained in this study are complementary to the cited results of the above authors. Consequently, both sets of data taken together represent a complete description of S20,* vs. % reactivity with CMC. It is shown that the model in which superhelical DNA is proposed to contain small intrastrand hairpin regions can be extended to account for the observed transitions in S20,* vs. reactivity.