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C D Collard

Publications and source records attributed to C D Collard.

28 records · Page 2Linked to original sources
PubMed1998

Complement activation following reoxygenation of hypoxic human endothelial cells: role of intracellular reactive oxygen species, NF-kappaB and new protein synthesis.

Complement plays an important role in ischemia-reperfusion injury. We recently demonstrated that reoxygenation of hypoxic human umbilical vein endothelial cells (HUVECs) activated the classical complement pathway and augmented iC3b deposition. In the present study, we investigated the potential role of oxygen-derived free radicals, NF-kappaB and new protein synthesis in this model. HUVECs subjected to 12 or 24 h hypoxic stress (1% O2) and then reoxygenated (0.5, 1, 2 or 3 h; 21% O2) in 30% human serum activated complement and deposited iC3b. Addition of hydrogen peroxide (H2O2; 1-100 micromol/l) to normoxic HUVECs increased iC3b deposition in a concentration-dependent manner. H2O2 (10 micromol/l), a concentration that did not significantly increase iC3b deposition on normoxic HUVECs, augmented iC3b deposition on hypoxic/reoxygenated HUVECs. We observed a significant increase in intracellular H2O2 and hydroxyl radical (OH.) production in hypoxic/reoxygenated HUVECs using dihydrorhodamine 123. Further, treatment of HUVECs with dimethylthiourea (DMTU, 1-100 micromol/l), deferoxamine (DEF, 1-100 micromol/l), or oxypurinol (10 micromol/l), but not superoxide dismutase (SOD, 500 U/ml), catalase (300 U/ml) or iron-loaded DEF, attenuated iC3b deposition following hypoxia/reoxygenation in a concentration-dependent manner. Western analysis demonstrated hypoxia-induced nuclear NF-kappaB translocation that increased with reoxygenation. Inhibition of new protein synthesis (i.e. cycloheximide) or inhibition of NF-kappaB (ALLN or SN-50) also significantly decreased iC3b deposition on hypoxic/reoxygenated HUVECs. We conclude that (1) hypoxic/reoxygenated HUVECs generate H2O2 and OH.; (2) treatment of HUVECs with cell permeable reactive oxygen species inhibitors/scavengers (i.e. DEF, DMTU, oxypurinol) but not large molecular weight inhibitors (i.e. catalase or SOD) significantly reduces iC3b deposition and (3) inhibition of new protein synthesis or NF-kappaB activation attenuates iC3b deposition. These data suggest that iC3b deposition on the vascular endothelium may be regulated by intracellular oxygen-derived free radical-induced activation of NF-kappaB, new protein synthesis and activation of the classical complement pathway during ischemia/reperfusion.

Cell Hypoxia↗
PubMed1997

Reoxygenation of hypoxic human umbilical vein endothelial cells activates the classic complement pathway.

BACKGROUND: Ischemia-reperfusion injury leads to the activation and endothelial deposition of complement. We investigated whether exposure of human umbilical vein endothelial cells (HUVECs) to hypoxia and/or reoxygenation activates complement and decreases HUVEC-surface expression of the C3 regulatory proteins CD46 and CD55. METHODS AND RESULTS: HUVECs were subjected to 0, 12, or 24 hours of hypoxia (O2 = 1%) and then reoxygenated for 3 hours (O2 = 21%) in the presence of 30% human serum. C3 deposition and HUVEC-surface expression of CD46 and CD55 were evaluated by ELISA and flow cytometry. C3 deposition on HUVECs subjected to 12 or 24 hours of hypoxia followed by 3 hours of reoxygenation was significantly greater than normoxic HUVECs. Inhibition of the classic but not the alternative complement pathway during reoxygenation attenuated C3 deposition. Western blot analysis of HUVEC lysates under reducing conditions demonstrated significantly increased iC3b deposition in hypoxic/reoxygenated HUVECs compared with normoxic HUVECs. FACS analysis confirmed iC3b deposition. HUVEC-surface expression of CD46 and CD55 increases after hypoxia and/or reoxygenation. CONCLUSIONS: We conclude that (1) hypoxia and reoxygenation of HUVECs significantly increases iC3b deposition on HUVECs, (2) C3 deposition after hypoxia and reoxygenation is largely mediated by the classic complement pathway, and (3) HUVEC-surface expression of CD46 and CD55 increases after hypoxia and reoxygenation. These data demonstrate that hypoxia and reoxygenation of human endothelial cells activates the classic complement pathway despite an increase in complement C3 regulatory proteins.

Antigens, CD↗
PubMed1996

Cumulative reduction in serum cholinesterase following repeated therapeutic plasma exchange.

STUDY OBJECTIVE: To investigate the magnitude of serum cholinesterase reduction following repeated therapeutic plasma exchange in patients with neuromuscular disease. DESIGN: Serum cholinesterase activity was measured immediately before and after each plasma exchange in open-label fashion and then analyzed using an analysis of variance model. SETTING: Inpatient neurology and allergy and immunology clinics at a university-affiliated hospital. PATIENTS: 50 consecutive patients with neuromuscular disease. INTERVENTIONS: All patients underwent repeated therapeutic plasma exchange, with each subject receiving up to a maximum of six plasma exchanges. MEASUREMENTS AND MAIN RESULTS: Serum cholinesterase activity was determined spectrophotometrically. Analysis of variance revealed a significant reduction in serum cholinesterase following each therapeutic plasma exchange (p < 0.0001), a significant and consistent reduction across the six treatments (p < 0.0001), and a significant interaction between (before versus after exchange) and treatment number (p < 0.0001). Mean serum cholinesterase before repeated therapeutic plasma exchange was 4817 U/L, but it decreased to a mean of 929 U/L following six plasma exchanges. CONCLUSIONS: There is a significant reduction in serum cholinesterase following repeated therapeutic plasma exchange. It is suggested that drugs metabolized by serum cholinesterase (e.g., succinylcholine, mivacurium) be used with caution in the period immediately following repeated therapeutic plasma exchange.

Anesthesia↗
PubMed1996

Complement-induced endothelial dysfunction in rabbits: mechanisms, recovery, and gender differences.

Activation of complement and attenuation of endothelium-dependent relaxation occur in a number of pathophysiological conditions. The aim of this study was to investigate the mechanisms of human complement activation and loss of endothelium-dependent relaxation in rabbit tissue, the duration of this loss, and the effects of gender and serum concentration. In rabbit thoracic aortic rings precontracted with phenylephrine, human serum (HS) concentration dependently induced a loss of endothelium-dependent relaxation to the receptor-dependent vasodilator acetylcholine (ACh) and receptor-independent vasodilator calcium ionophore A23187. Serum-induced loss of ACh-dependent relaxation was decreased when rings were bathed in 1) HS depleted of factor B, C2, or C8, 2) heat-inactivated HS, or 3) HS with complement inhibitor sCR1 or sCR1[desLHR-A]. Superoxide dismutase had no effect on serum-induced loss of ACh-dependent relaxation. Serum-induced loss of ACh-dependent relaxation returned to control values after removal of HS. Serum-induced loss of ACh-dependent relaxation was greater in male than in female aortic rings. These results suggest that 1) complement activation directly attenuates endothelium-dependent relaxation via the classical and alternative pathways independent of superoxide anion formation, 2) this attenuation is concentration dependent, reversible, and dependent on formation of C5b-9, and 3) endothelial tissue from males is more susceptible than that from females to the acute effects of complement activation.

Acetylcholine↗
PubMed1989

Low current electrostimulation produces naloxone-reversible analgesia in rats.

A new form of transcranial electrostimulation (TE) has been shown to induce analgesia in rats, as measured by the wet tail flick test. Charge-balanced rectangular current pulses of very low amplitude were delivered bilaterally into low impedance regions of the rat pinnae. The resultant analgesia was studied as a function of systematic variations in stimulus frequency, amplitude and duration. The optimal current for inducing analgesia was found to be 10 microA, well below the startle threshold, and several orders of magnitude below effective stimulus current levels required for other treatment modalities. The optimal stimulation duration was 30 min, during which time a slow onset of analgesia was noted. Significant analgesia persisted for at least 200 min after stimulation ended, and no evidence was found of tolerance developing in the course of 5 daily stimulation sessions. Consistent with findings for other forms of electrostimulation, the analgesic effect of TE was abolished by subcutaneous injection of 3 mg/kg naloxone, suggesting that the mechanism of TE analgesia has an endogenous opioid component.

Animals↗
PubMedDate not supplied

Complement activation following oxidative stress.

It is clear that complement plays an important role in the inflammatory process following oxidative stress in cellular and animal models. Clinical trials underway with novel complement inhibitors will establish the potential therapeutic benefit of complement inhibition in human disease. For as much as we understand about the role of complement in disease states, many questions remain. How is complement activated on endothelial cells following oxidative stress? What is the ligand for MBL on endothelial cells following oxidative stress? Will inhibition of MBL provide tissue protection to the extent observed with other complement inhibitors such as sCR1 or anti-C5 mAbs? These questions and more will undoubtedly be answered in the next millennium.

Animals↗
PubMedDate not supplied

Pharmacology and biological efficacy of a recombinant, humanized, single-chain antibody C5 complement inhibitor in patients undergoing coronary artery bypass graft surgery with cardiopulmonary bypass.

BACKGROUND: Cardiopulmonary bypass (CPB) induces a systemic inflammatory response that causes substantial clinical morbidity. Activation of complement during CPB contributes significantly to this inflammatory process. We examined the capability of a novel therapeutic complement inhibitor to prevent pathological complement activation and tissue injury in patients undergoing CPB. METHODS AND RESULTS: A humanized, recombinant, single-chain antibody specific for human C5, h5G1.1-scFv, was intravenously administered in 1 of 4 doses ranging from 0.2 to 2.0 mg/kg before CPB. h5G1.1-scFv was found to be safe and well tolerated. Pharmacokinetic analysis revealed a sustained half-life from 7.0 to 14.5 hours. Pharmacodynamic analysis demonstrated significant dose-dependent inhibition of complement hemolytic activity for up to 14 hours at 2 mg/kg. The generation of proinflammatory complement byproducts (sC5b-9) was effectively inhibited in a dose-dependent fashion. Leukocyte activation, as measured by surface expression of CD11b, was reduced (P<0.05) in patients who received 1 and 2 mg/kg. There was a 40% reduction in myocardial injury (creatine kinase-MB release, P=0.05) in patients who received 2 mg/kg. Sequential Mini-Mental State Examinations (MMSE) demonstrated an 80% reduction in new cognitive deficits (P<0.05) in patients treated with 2 mg/kg. Finally, there was a 1-U reduction in postoperative blood loss (P<0. 05) in patients who received 1 or 2 mg/kg. CONCLUSIONS: A single-chain antibody specific for human C5 is a safe and effective inhibitor of pathological complement activation in patients undergoing CPB. In addition to significantly reducing sC5b-9 formation and leukocyte CD11b expression, C5 inhibition significantly attenuates postoperative myocardial injury, cognitive deficits, and blood loss. These data suggest that C5 inhibition may represent a novel therapeutic strategy for preventing complement-mediated inflammation and tissue injury.

Antibodies, Monoclonal↗