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Biomedical subjects

P C Giclas

Publications and source records attributed to P C Giclas.

At least 19 recordsLinked to original sources
PubMed2004

Increased serum C3 levels in Crry transgenic mice partially abrogates its complement inhibitory effects.

Complement receptor 1-related gene/protein y (Crry) is a potent murine complement regulator that inhibits C3 convertases. Transgenic mice that overexpress soluble Crry (sCrry), directed systemically by the metallothionein-I promoter, have been used as an animal model for chronic blockade of complement activation. Recently we have found that alternative pathway (AP) activity in Crry transgenic mice was not inhibited as much as expected. To elucidate the mechanism of this effect, we evaluated the AP activities and levels of sCrry and AP complement components in transgenic and non-transgenic mice. In transgenic mice, expression of sCrry was induced by feeding zinc sulphate solution to 70.1 +/- 42.7 micro g/ml mean serum level. Its corresponding level of purified sCrry inhibited 49% of AP activity of normal mice serum; however, the actual AP activities in transgenic mice were not decreased when compared to non-transgenic mice (130.2 +/- 9.0%versus 113.0 +/- 35.4%). Expressed sCrry was functional, as immunoprecipitation and removal of sCrry from transgenic sera with rabbit anti-Crry polyclonal antibody resulted in enhanced AP activity, consistent with initial levels of sCrry. We then compared the changes to C3, factor B, factor H and factor D serum levels in transgenic and non-transgenic mice after induction of sCrry expression. Of these only C3 was increased after zinc feeding in transgenic mice compared to non-transgenic mice (142.8 +/- 14.1%versus 121.4 +/- 15.1%, P = 0.023). These results suggest that the inhibitory effect of chronic exposure to sCrry is compensated by concomitant alteration in C3 levels. This result also suggests the presence of a complement regulatory protein controls the level of serum C3, which has potential importance in the design and interpretation of studies involving chronic use of complement inhibitors.

Animals↗
PubMed2003

Hybridomas expressing gammadelta T-cell receptors respond to cardiolipin and beta2-glycoprotein 1 (apolipoprotein H).

Hybridomas expressing murine gammadelta T-cell receptors were found to produce cytokines in response to cardiolipin (CL) and structurally related anionic phospholipids. This response required serum at concentrations related to the amount of CL in cultures. The purified serum factor, beta2-glycoprotein 1 (beta2-GP1) (apolipoprotein H), supported the CL response alone, whereas several other serum proteins and ovalbumin did not. beta2-GP1 is known to form complexes with anionic phospholipids, particularly CL, which are often recognized by pathological autoantibodies. We speculate that gammadelta T cells also recognize such complexes and that the hybridoma response reported here reflects this specificity.

Animals↗
PubMed2000

A comparative study of mammalian and reptilian alternative pathway of complement-mediated killing of the Lyme disease spirochete (Borrelia burgdorferi).

The potential bactericidal activity of the alternative complement pathway of mammalian and reptilian sera to Borrelia burgdorferi sensu stricto (s.s.) was evaluated in vitro. Complement-mediated killing was observed when cultured spirochetes were inoculated into sera from the western fence lizard (Sceloporus occidentalis) and from the southern alligator lizard (Elgaria multicarinata), but not when they were inoculated into serum from either the deer mouse (Peromyscus maniculatus) or from humans. Spirochetes were still alive after 4 hr in lizard serum that had been preheated at 56 C for 30 min to inactivate complement. Furthermore, when lizard serum was chelated with 10 mM ethylenediaminetetraacetic acid to block all complement activation, borreliacidal activity was arrested. When lizard serum was chelated with 10 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid plus 4 mM MgCl2 to block only classical complement pathway activation, >85% of spirochetes were immobilized within 1 hr. Differences in B. burgdorferi s.s. mortality were not observed when chelators with or without MgCl2 were added to serum from either deer mice or humans. Proteins comprising the alternative complement pathway are responsible for the borreliacidal activity observed in the blood of S. occidentalis and E. multicarinata.

Animals↗
PubMed1997

Rush immunotherapy results in allergen-specific alterations in lymphocyte function and interferon-gamma production in CD4+ T cells.

BACKGROUND: Allergen immunotherapy results in a number of changes in clinical, inflammatory, and immunologic parameters. However, the basis for the specificity of this form of therapy is unknown, especially in the context of changes in T- and B-lymphocyte function after desensitization to specific allergens. OBJECTIVE: This study was designed to determine the immunologic consequences of rush immunotherapy. METHODS: We studied 10 patients who had positive skin test responses to the house dust mite Dermatophagoides pteronyssinus (Dpt) and cat dander extract. Each received rush immunotherapy to mite, but not cat dander, over a 2- to 4-week period until maintenance was achieved. Patients were evaluated before and when maintenance was achieved for skin test and nasal reactivity to mite and cat dander; antibody levels to the allergen were monitored, as were lymphocyte proliferative responses and cytokine production. RESULTS: Rush immunotherapy to house dust mite resulted in a significant reduction in skin and nasal reactivity to mite allergen, but not to cat allergen, in 10 of 10 patients. This was accompanied by a rise in serum anti-Dpt IgE, whereas anti-cat IgE was not altered (7 of 7 patients). In seven of seven patients there was an increase in anti-Dpt IgG4 levels. T-cell proliferative responses to mite antigen were suppressed, and numbers of CD8+ T cells increased in frequency. There was a marked increase in interferon-gamma production, particularly by CD4+ T cells in 10 of 10 patients. The correlation between the increases in interferon-gamma production and the changes in cutaneous reactivity was highly significant. CONCLUSION: We show that rush immunotherapy is immunologically specific in eliciting changes in T- and B-cell responses to the desensitization antigen. The specificity and potential benefit of immunotherapy may be linked to the increase in interferon-gamma production by allergen-activated CD4+ T lymphocytes.

Adolescent↗
PubMed1997

Age-dependent neutrophil and blood flow responsiveness in acute pulmonary inflammation in rabbits.

Diminished ability of neonatal neutrophils to orient and move in a chemotactic gradient has been linked to compromised pulmonary host defense. We investigated whether deficiency of neonatal neutrophil function in vitro was evident in acute pulmonary inflammation. Analysis of neutrophils in vitro showed impaired chemotaxis in 4-wk-old compared with adult rabbits. In vivo-directed migration of labeled neutrophils into the alveolar space of adult rabbits in response to C5f instillation was significantly less for neutrophils donated from 4-wk-old rabbits compared with those from adults. In contrast, there were no differences in the alveolar accumulation of 4-wk-old and adult labeled neutrophils in 4-wk-old rabbits in response to C5f instillation, although the response showed a shorter time course than seen in adult rabbits. Adult rabbits diverted 46% of the blood away from the right cranial lung lobe, whereas 4-wk-old rabbits showed no change in blood flow after C5f instillation. Megakaryocytes (a source of blood flow mediators) were 3.2-fold greater in adult compared with 4-wk-old lung. These data suggest that the lack of blood flow diversion from inflamed neonatal lung increases neutrophil migration into alveoli, allowing for preservation of an inflammatory response despite neutrophil deficiencies in chemotaxis.

Acute Disease↗
PubMed1997

Serum ferritin as a predictor of the acute respiratory distress syndrome.

We investigated serum ferritin levels as a predictor of the acute respiratory distress syndrome (ARDS) because: (1) proinflammatory cytokines, which are implicated in ARDS, increase ferritin synthesis; and (2) oxidative stress in patients at risk for ARDS might liberate iron from ferritin, accelerating toxic hydroxyl radical (.OH) formation. Serum ferritin levels measured by radioimmunoassay (RIA) were greater in 75 patients at risk for ARDS (women, p < 0.0001; men, p < 0.0001) and 8 patients with ARDS (women, p = 0.001; men, p = 0.0009) than in healthy control subjects. Serum ferritin levels were also greater in female (p = 0.003) and male (p = 0.003) at-risk patients who developed ARDS than in patients who did not develop ARDS. In women, a value exceeding 270 ng/ml predicted ARDS with an 83% sensitivity, 71% specificity, 67% positive, and 86% negative predictive value. In men, a value exceeding 680 ng/ml predicted ARDS with a 60% sensitivity, 90% specificity, 75% positive, and 82% negative predictive value. Serum ferritin levels did not correlate with C-reactive protein levels, were not different in medical or surgical at-risk patients, and were not accounted for by liver disease. Evaluating serum ferritin levels in at-risk patients may help predict the development of ARDS and thereby improve study and treatment of ARDS. Elevated serum ferritin levels may also regulate the participation of iron in the oxidative responses that contribute to ARDS.

Adult↗
PubMed1997

Activation of complement by tissue plasminogen activator, but not acute cerebral ischemia, in a rabbit model of thromboembolic stroke.

Although complement activation is associated with tissue injury during inflammatory and ischemic states, complement activation in states of acute cerebral ischemia before and after administration of tissue plasminogen activator (TPA) has not yet been examined and is the focus of this investigation. Twenty-four New Zealand White rabbits weighing 3 to 3.5 kg were used for this study. Of these, 20 were subjected to intracranial autologous clot embolization via the internal carotid artery. Three hours postembolization, rabbits received an intravenous infusion of TPA (6.3 mg/kg, 20% bolus with the remainder infused over a 2-hour interval; 12 animals) or vehicle (eight animals). All animals were observed for a total of 7 or 8 hours postembolization. These two groups were compared to a cohort undergoing sham operation with subsequent TPA infusion (four animals). Plasma samples to quantify complement component C5 hemolytic activity (C5H5O) were obtained at the following time points: 30 minutes before and after clot embolization; 1 hour before and 1 hour after the initiation of therapy with TPA or vehicle and at the completion of the protocol; 7 to 8 hours after clot embolization. The C5 activation was not detected as the result of acute cerebral ischemia. However, animals receiving TPA with or without concomitant clot embolization exhibited C5 activation as assessed by a reduction in C5 hemolytic function, both 1 hour after initiation of TPA infusion (78.7 +/- 10.3% and 77.5 +/- 9.9% of baseline value, respectively; mean +/- standard error of the mean [SEM]) and at the end of the protocol, 2 hours after the completion of the TPA infusion (72.5 +/- 8.8% and 53.3 +/- 8.1%, respectively; mean +/- SEM, p < 0.05, each group). This study supports the conclusion that TPA, but not acute cerebral ischemia, may activate the complement cascade in this rabbit model of thromboembolic stroke.

Animals↗
PubMed1997

Activation of the alternative pathway of complement by a phosphorothioate oligonucleotide: potential mechanism of action.

Intravenous infusion of high doses of phosphorothioate oligonucleotides in monkeys has been associated with transient alterations in hematologic and hemodynamic parameters, which appear to be secondary to complement activation. ISIS 2302, a phosphorothioate oligonucleotide specific for human intracellular adhesion molecule-1, was used to further characterize complement activation in monkeys. Complement activation occurred selectively through the alternative pathway resulting in increased plasma concentrations of the complement split products Bb, C3a and C5a. Marked fluctuations in circulating neutrophil counts and reductions in cardiac output were closely associated with peak production of anaphylatoxins C3a and C5a. Changing both dose and infusion duration revealed that complement activation is related to plasma levels of oligonucleotide, and that there is a minimum threshold concentration of approximately 50 micrograms/ml of ISIS 2302 that is required to activate complement. Dose regimens in which plasma concentrations do not exceed this threshold do not result in complement activation. Further investigation reveals that plasma concentrations of a key regulatory component of the alternative pathway, Factor H, were also decreased after administration of ISIS 2302. Decreases in Factor H levels are suggestive of a possible mechanism of complement activation. Direct interaction between ISIS 2302 and Factor H was demonstrated in a competition assay, where increasing concentrations of ISIS 2302 eluted Factor H from a heparin-sepharose column. These data demonstrate a clear correlation between plasma oligonucleotide concentrations and complement activation. Interactions between ISIS 2302 and Factor H may lead to activation of the alternative complement pathway.

Animals↗
PubMed1996

Invasive Haemophilus influenzae type b infection in a child with familial deficiency of the beta subunit of the eighth component of complement.

A child who had had meningitis caused by Haemophilus influenzae type b, and then had meningococcal meningitis, was found to have familial deficiency of the beta subunit of the eighth component of complement. The child had not received the H. influenzae type b vaccine. If this deficiency is discovered, we recommend that family members be screened, regardless of their health status.

Complement C8↗
PubMed1994

Complement activation and hemodynamic changes following intravenous administration of phosphorothioate oligonucleotides in the monkey.

Rapid intravenous infusion of GEM 91, a 25-mer phosphorothioate oligonucleotide complementary to the gag site of HIV, in the monkey produces transient decreases in peripheral total WBC and neutrophil counts, hemoconcentration, and a brief increase followed by a prolonged decrease in arterial blood pressure. These changes are preceded by and are likely mediated by activation of C5 complement. These effects are dose and infusion rate dependent and can be avoided by administering GEM 91 by slow intravenous infusion. Similar hemodynamic effects are produced with rapid intravenous infusion of other phosphorothioate oligonucleotides varying in length from 20- to 33-mer, and are, therefore, not sequence specific but a property of this chemical structure.

Animals↗
PubMed1993

Peripheral bypass-induced pulmonary and coronary vascular injury. Association with increased levels of tumor necrosis factor.

BACKGROUND: Although cardiopulmonary bypass is associated with systemic complement activation and neutrophil sequestration, it is unclear whether bypass-induced vascular injury is localized and dependent on organ ischemia. We hypothesized that other factors perhaps related to placement of a bypass circuit or to blood perfusion of a pump-oxygenator system may produce vascular injury caused by systemically circulating mediators. In dogs, we determined whether application of a systemic venoarterial bypass circuit with pump-oxygenator perfusion but without pulmonary or cardiac flow diversion (peripheral bypass) leads to vascular injury. Since several features of the postperfusion syndrome after bypass resemble sequelae of endotoxin exposure, we also measured circulating endotoxin and tumor necrosis factor levels. METHODS AND RESULTS: Anesthetized dogs underwent 2 hours of exposure to a pump-oxygenator with peripheral venoarterial bypass. We used a double indicator measurement of pulmonary and coronary vascular permeability (protein leak index [PLI]) as indexes of vascular injury. Compared with controls (n = 7), the pulmonary PLI of dogs undergoing bypass (n = 11) increased more than threefold (18.8 +/- 2.3 vs 63.3 +/- 7.6 x 10(-4) min-1; P < .05) and the coronary PLI increased more than twofold (P < .05). The rate of disappearance of intravascular radiolabeled protein increased threefold after bypass (disappearance t1/2, 241 +/- 35 vs 84 +/- 15 minutes, control vs bypass; P < .05), suggesting a generalized increase in vascular permeability. Circulating endotoxin was detectable in blood samples from 8 of 8 bypass animals (range, 0.24 to 4.56 ng/mL) compared with 2 of 5 controls (P < .05). Tumor necrosis factor levels increased significantly with bypass (6.7 +/- 3.8 vs 146.7 +/- 33.6 U/mL, baseline vs bypass; P < .05) and were only slightly and nonsignificantly increased in controls (7.0 +/- 4.4 vs 18.2 +/- 5.9 U/mL; P = NS). Peak tumor necrosis factor but not peak endotoxin levels correlated with pulmonary and with coronary protein leak. As expected, circulating complement (CH50) levels decreased significantly during bypass, reflecting systemic complement activation. However, the levels correlated poorly with the severity of vascular injury. CONCLUSIONS: We conclude that peripheral pump-oxygenator bypass causes coronary and pulmonary vascular injury that is independent of blood flow diversion and is associated with the appearance of circulating levels of endotoxin and tumor necrosis factor, which may play a role in bypass-induced vascular injury.

Animals↗
PubMed1991

The effect of intravascular complement activation and brief episodes of hypoxia on protein in bronchoalveolar lavage fluid in C5 sufficient and deficient mice.

Our earlier investigations indicated that systemic complement activation with iv cobra venom factor (CVF) or infused zymosan-activated rabbit plasma or rabbit C5a does not significantly increase bronchoalveolar lavage albumin in rabbits (Am Rev Respir Dis 1982; 125:335-340); but that complement activation due to CVF combined with a brief episode of hypoxia increases lavage albumin and is associated with the presence of neutrophils for its expression (J Clin Invest 1985; 75:902-910). In order to determine if intravenous CVF and hypoxia cause similar alterations in mice, and to investigate the time course of the response as well as the importance of C5 fragments to the process, we challenged the B10.D2/nSn strain of C5 sufficient mice (C5+) and the congenic B10.D2/oSn strain of C5 deficient mice (C5-) with intravenous CVF, 15 min of 12% oxygen, or CVF followed by hypoxia. Neither C5+ nor C5- mice had significant increases in lavage protein after either CVF or hypoxia. However, the combined insults significantly increased lavage protein in C5+ but not C5- mice; polyacrylamide gel electrophoresis showed increased amounts of proteins of low and high molecular weights in lavage fluid from the C5+ strain. While the time course of abnormalities in mice was different from that in rabbits, both meclofenamate pretreatment and neutrophil depletion attenuated the increases in lavage protein after the combined insults in both animal species. Infusion of prostaglandin E2 (PGE2) with CVF in the C5+ mice also led to significant increases in lavage protein. We conclude that in mice, intravenous complement activation, as an isolated event, does not cause a significant increase in lavage protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Complement activation is a secondary rather than a causative factor in rabbit pulmonary artery ischemia/reperfusion injury.

We have previously demonstrated that reperfusion of a rabbit lung in vivo after 24 h of unilateral pulmonary artery occlusion results in edema, transient leukopenia, and intravascular leukocyte aggregation. We hypothesized that complement was activated by reperfusion and that this in turn contributed to lung injury. In the preliminary phase of the study, we found that ischemia followed by reperfusion resulted in a drop in C3 to 15 +/- 10% (mean +/- SEM) of the prereperfusion value as compared with no change in a group of control animals that had undergone an identical thoracotomy but without pulmonary artery occlusion and reperfusion (p less than 0.05). We then studied three groups of animals to determine if complement depletion with cobra venom factor (CVF) prior to ischemia and reperfusion would prevent the injury. Rabbits treated with CVF but without occlusion and reperfusion did not develop significant lung edema, with left and right lung wet/dry ratios of 5.32 +/- 0.11 and 5.26 +/- 0.12, respectively. For rabbits that were not treated with CVF but underwent ischemia and reperfusion, the comparable numbers were 6.15 +/- 0.36 and 5.19 +/- 0.32 (p less than 0.05 for right versus left). For CVF-treated rabbits that underwent ischemia and reperfusion, the right/left difference persisted (6.77 +/- 0.48 versus 5.35 +/- 0.14, p less than 0.01). Immunocytochemistry documented C3 deposition in non-CVF rabbits that underwent ischemia and reperfusion but not in CVF-treated rabbits. We conclude that ischemia/reperfusion of the lung results in complement activation, but it is not a complement-dependent injury.

Animals↗
PubMed1990

The terminal complement complex (sC5b-9) is not specifically associated with the development of the adult respiratory distress syndrome.

Previous investigators suggested that increased plasma levels of the terminal complement complex (sC5b-9) are an early marker for the development of adult respiratory distress syndrome (ARDS) in septic patients. We asked whether an increase in sC5b-9 was also associated with the development of ARDS from other etiologies and whether sC5b-9 measurements consistently reflected complement activation in vivo. We evaluated 75 patients with sepsis, trauma, hypertransfusion, multiple fractures, aspiration, or pancreatitis who were at risk for ARDS but did not develop the syndrome and 23 patients with similar histories who did develop ARDS. Of the latter patients, seven were identified and studied both when they were at risk and when they had ARDS. Serial blood samples were obtained and analyzed for the complement activation products Bb, Ba, C4d, C3d, IC3b, and sC5b-9. All but one of the patients studied had levels of one or more complement fragments that were greater than 2 SD above the mean obtained from 18 normal subjects. In contrast to the report referred to previously, none of the fragments measured, including sC5b-9, was a specific indicator of ARDS, and no combination of complement fragments predicted which patients at risk would develop ARDS. Patients demonstrated evidence of activation of the classical pathway only, alternative pathway only, or both pathways, but none of these was associated with greater risk or severity of disease. In addition, in several patients only late components were activated, suggesting that enzymes other than those derived from complement activation may be responsible. In conclusion, complement can be activated by a variety of mechanisms in critically ill patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Complement Activation↗
PubMed1989

Cessation of neutrophil influx in C5a-induced acute experimental arthritis is associated with loss of chemoattractant activity from the joint space.

Neutrophil emigration is a critical component of the inflammatory process and is generally thought to play a role in host defense as well as in the tissue injury that often accompanies inflammation. Most inflammatory reactions exhibit a sequence of emigrating cell types, thus clearly demonstrating that the neutrophil influx eventually ceases and that the neutrophils are then removed from the lesion. It has been our premise that in order to understand the processes that lead to the progressive inflammatory reactions that underly so many disease processes, it is important to determine the mechanism by which the "normal" inflammatory response resolves. The purpose of this study was to identify the time of cessation of neutrophil influx in experimental arthritis induced by the injection of C5 fragments (C5f) and to investigate mechanisms underlying the cessation process. The migration of i.v. delivery pulses to inflamed joints was assessed by lavage of the joint space and by external scintigraphy. We found no evidence for the development of inhibitory systems against chemotactic factors or "desensitization" of the inflamed site, because a second injection of C5f into joints which had been injected previously with C5f resulted in enhancement rather than inhibition of migration. Neither was evidence found for altered tissue barriers to migration or for desensitization of neutrophils as possible explanations for cessation of influx. The major mechanism appeared to be a loss of chemoattractant activity in the joint space between 2 h and 6 h after C5f injection which was detected by transfer into a fresh joint. Radiolabeled C5a des-Arg had a t1/2 of disappearance from the joint of less than 1 h, which suggested that the transferred chemoattractant must, in part, have been due to the generation of new chemotaxins by C5f injection. These observations suggest that continued generation of chemoattractants or failure of their subsequent removal may be mechanisms leading to persistent neutrophil influx in chronic inflammation.

Acute Disease↗
PubMed1989

Absence of induction of IL-1 production in human monocytes by complement fragments.

The ability of C fragments to induce IL-1 production in human monocytes was examined by using various approaches to carefully exclude the role of contaminating endotoxin. The presence of IL-1 activity in monocyte supernatants and lysates was assayed by the augmentation of PHA-induced proliferation of murine thymocytes. SRBC were opsonized with IgM rabbit antibodies and various human C components to prepare EAC reagents that contained less than 25 pg LPS/ml of EAC at 5 x 10(8) cells/ml. EAC1q, EAC4b, EAC4b2aoxy, EAC4b2aoxy C3b, EAC4b2aoxyC3bi, and EAC4b2aoxyC3d all failed to induce IL-1 production when incubated at 10- to 100-fold excess with adherent human monocytes. Similarly, LPS-free purified C3a, C5a, and C5a des Arg all showed no IL-1-inducing activities at concentrations up to 25 micrograms/ml. However, the same C5a preparations were active on human monocytes in the induction of chemotaxis, and C3a and C5a both induced skin-blueing in guinea pigs. Fragment Ba and Bb preparations purified by gel filtration chromatography contained approximately 100 pg LPS/micrograms Ba or Bb. These Ba and Bb preparations at 10 and 50 micrograms/ml, respectively, induced IL-1 production in the presence of 5 micrograms/ml polymyxin B (PMB). However, Ba and Bb preparations purified by affinity chromatography and HPLC contained lower levels of endotoxin contamination and displayed IL-1-inducing activities at Ba and Bb concentrations of 50 and 100 micrograms/ml, respectively, that were almost completely inhibited by PMB. To explore further the role of contaminating endotoxin, a Bb preparation was adsorbed with PMB-4B in the presence of a dialyzable detergent to remove LPS bound to the Bb. This LPS-free Bb preparation failed to induce IL-1 production while maintaining intact enzymatic activities. These results indicate that various solid phase or soluble C fragments, including C3b, iC3b, C3d, C3a, C5a, Ba or Bb do not induce IL-1 production in human monocytes in the absence of contaminating endotoxin.

Antigen-Antibody Complex↗