Biomedical subjects

G O Till

Publications and source records attributed to G O Till.

At least 19 recordsLinked to original sources

PubMed1998

Partial liquid ventilation decreases albumin leak in the setting of acute lung injury.

PURPOSE: This study evaluated the ability of partial liquid ventilation (PLV, gas ventilation of the perfluorocarbon-filled lungs) to reduce the amount of lung albumin leak present in the setting of acute lung injury. MATERIALS AND METHODS: An experimental controlled, randomized design was used. All studies were performed in the liquid ventilation laboratories at the University of Michigan Medical Center. Twenty-five Sprague-Dawley male rats 500+/-50 g were divided into five experimental groups: (1) CVF only (n=5), animals were cobra venom factor (CVF) lung injured; (2) PLV-CVF (n=5) animals received perflubron and PLV before CVF lung injury; (3) CVF-PLV (n=5) animals received PLV after CVF lung injury; (4) PLV only (n=5) animals underwent partial liquid ventilation without lung injury; and (5) Gas only (n=5) animals underwent gas ventilation without lung injury. In all groups iodinated bovine serum albumin (125I-BSA) was delivered by intravenous injection along with CVF or a saline placebo. RESULTS: When the CVF animals were compared with all other groups, a decrease in albumin leak was observed for all groups when compared with the CVF only controls (P < .001 by ANOVA; CVF only=1.22+/-0.12 versus PLV-CVF=0.46+/-0.08, P < .001; CVF-PLV=0.70+/-0.25, P < .001; PLV only=0.22+/-0.01, P < .001; Gas only=0.17+/-0.02, P < .001). CONCLUSIONS: These data suggest that intratracheal instillation of perfluorocarbon before or after induction of lung injury results in a reduction in pulmonary albumin leak.

Analysis of Variance↗

PubMed1998

Mediators of microvascular injury in dermal burn wounds.

In previous studies we have demonstrated that second-degree thermal injury of skin in rats leads to secondary effects, such as systemic complement activation, C5a-mediated activation of blood neutrophils, their adhesion-molecule-guided accumulation in lung capillaries and the development of acute pulmonary injury, largely caused by neutrophil-derived toxic oxygen metabolites. In the dermal burn wound, however, pathophysiologic events are less well understood. The injury is fully developed at four hours post-burn. To further elucidate the pathogenesis of the "late phase" dermal vascular damage, rats were depleted of neutrophils or complement by pretreatment with rabbit antibody against rat neutrophils or with cobra venom factor, respectively. In other experiments, rats were treated with blocking antibodies to IL-6, IL-1, and TNF alpha immediately following thermal burning or were pretreated with hydroxyl radical scavengers (dimethyl sulfoxide, dimethyl thiourea). Extravasation of 125I-labeled bovine serum albumin into the burned skin was studied, as well as, skin myeloperoxidase levels. The studies revealed that, like in secondary lung injury, neutrophils and toxic oxygen metabolites, are required for full development of microvascular injury. In contrast, however, development of dermal vascular damage in thermally injured rats was not affected by complement depletion. Our data suggest that the development of microvascular injury in the dermal burn wound is complement-independent, involves the pro-inflammatory cytokines IL-1, TNF alpha and IL-6, and may result from reactive oxygen metabolites generated by neutrophils accumulating in the burn wound.

Animals↗

PubMed1998

Neutrophil accumulation is reduced during partial liquid ventilation.

OBJECTIVE: This study evaluates the ability of perflubron to inhibit pulmonary neutrophil accumulation during partial liquid ventilation (PLV) in the setting of acute lung injury. DESIGN: Randomized, controlled, nonblinded study. SETTING: Research laboratory at a university. SUBJECTS: Male, Sprague-Dawley rats (n = 120, 506 +/- 42 g). INTERVENTIONS: Animals were divided into eight groups (n = 15 in each group, of which n = 12 for myeloperoxidase content and n = 3 for histologic neutrophil counting): a) GV-CVF group, animals received gas ventilation (GV) with the induction of lung injury using cobra venom factor (CVF); b) PLV-CVF group, animals received partial liquid ventilation before the induction of lung injury; c) PEEP-CVF group, animals received positive end-expiratory pressure (PEEP) before the administration of cobra venom factor; d) CVF-PLV group, animals received partial liquid ventilation after cobra venom factor; e) CVF-PEEP group, animals received PEEP after cobra venom factor; f) PLV only group, animals received partial liquid ventilation only; g) GV only group, animals received gas ventilation only; and h) NVSBA group, nonventilated spontaneous breathing animals. MEASUREMENTS AND MAIN RESULTS: After the experimental period, total lung myeloperoxidase content was significantly decreased in the PLV-CVF (0.29 +/- 0.08, p = .02) and PEEP-CVF (0.34 +/- 0.04, p = .01) groups when compared with the GV-CVF group (0.62 +/- 0.07). When compared with the GV-CVF group, a trend toward a reduction in myeloperoxidase was observed in the CVF-PLV (0.42 +/- 0.05, p = .07) and the CVF-PEEP (0.39 +/- 0.06, p = .07) groups. When compared with the cobra venom factor only group (GV-CVF 47 +/- 2 neutrophils/high-power field), reductions in neutrophil count were observed in all groups (neutrophils/high-power field): PLV-CVF (20 +/- 2, p = .009); PEEP-CVF (24 +/- 1, p = .01); CVF-PLV (30 +/- 2, p = .03); and CVF-PEEP (37 +/- 1, p = .04). CONCLUSION: These data suggest that both partial liquid ventilation and PEEP result in a reduction in neutrophil accumulation in the setting of acute lung injury.

Animals↗

PubMed1998

The pattern of early lung parenchymal and air space injury following acute blood loss.

UNLABELLED: Acute lung injury is a frequent clinical occurrence following blood loss and trauma. The nature of this injury remains poorly understood. OBJECTIVE: To examine the relative parenchymal and intra-alveolar distribution of inflammation in a rat model of hemorrhage and resuscitation. METHODS: Rats were anesthetized and subjected to hemorrhage followed by resuscitation with shed blood and saline. Myeloperoxidase activity of lung homogenates and cytology of bronchoalveolar lavage fluid were used to measure total lung and intra-alveolar neutrophil invasion. Extravasation of i.v.-administered [125I]-albumin was used to determine total lung and alveolar permeability. Permeability results were analyzed using their base-10 logarithmic transformations. RESULTS: 86 animals were studied. Whole-lung myeloperoxidase activity was increased (control = 0.34 +/- 0.16 units, injured = 0.84 +/- 0.43 units, p < 0.01), while there was no difference in intra-alveolar leukocyte counts (injured = 1.85 +/- 1.30 x 10(5)/mL, control = 2.44 +/- 1.75 x 10(5)/mL, p = 0.40), suggesting that the cellular component of the injury was more severe in the intravascular and interstitial spaces. There was a strong trend toward increased permeability in the interstitial compartment, and a significant increase in permeability in the intra-alveolar compartment (whole-lung permeability: control = -0.27 +/- 0.19 units, injured = 0.10 +/- 0.55 units, p = 0.06; alveolar permeability: control = -2.00 +/- 0.47 units, injured = -1.32 +/- 0.49 units, p < 0.01), suggesting that the loss of integrity to macromolecules was not limited to the interstitium. CONCLUSION: Hemorrhage and resuscitation resulted in an acute lung injury characterized by extravasation of intravascular protein into both the interstitium and the intra-alveolar space. Neutrophil invasion of the lung was demonstrable only in the interstitial compartment.

Animals↗

PubMed1997

A major role for neutrophils in experimental bullous pemphigoid.

Bullous pemphigoid (BP) is an inflammatory subepidermal blistering disease associated with an IgG autoimmune response to the hemidesmosomal protein, BP180. Using a passive transfer mouse model, our group has shown previously that antibodies to the murine BP180 (mBP180) ectodomain are capable of triggering a blistering skin disease that closely mimics human BP. In this study, we investigated the role of neutrophils in the immunopathogenesis of this disease model. BALB/c mice depleted of circulating neutrophils by treatment with neutrophil-specific antibodies were no longer susceptible to the pathogenic effects of anti-mBP180 IgG. IgG and complement were deposited at the dermal-epidermal junction of these animals, but there was no evidence of inflammatory infiltration or blistering. C5-deficient mice, which are resistant to the pathogenic activity of anti-mBP180 IgG, could be made susceptible to this IgG-mediated blistering disease by intradermal administration of a neutrophil chemoattractant, IL-8 or C5a. Intraperitoneal injection of IL-8, which sequesters neutrophils in the peritoneal cavity, interferes with anti-mBP180-induced neutrophilic infiltration of the skin and prevented the development of BP disease in BALB/c mice. These findings provide the first direct evidence that neutrophils recruited to the skin via a C5-dependent pathway play an essential role in subepidermal blister formation in experimental BP, and suggest new directions for disease intervention.

Animals↗

PubMed1997

C5a-dependent up-regulation in vivo of lung vascular P-selectin.

Acute lung injury in rats following systemic activation of complement by i.v. infusion of cobra venom factor (CVF) is known to be P-selectin dependent. In the current studies, infusion of CVF caused the appearance in plasma of C5a (as revealed by ELISA analysis) together with neutrophil chemotactic activity, which was totally blocked by addition in vitro of anti-rat C5a. Using a detector 125I-labeled antibody to rat P-selectin, we have demonstrated quantitative up-regulation in vivo of lung vascular P-selectin in a time-dependent manner after infusion of CVF. This up-regulation was almost completely blocked by prior complement depletion or by the infusion of anti-rat C5a. Platelet depletion did not affect CVF-induced up-regulation of lung vascular P-selectin, indicating that platelets were not the source of P-selectin. These results demonstrate that complement and, specifically, C5a are necessary for up-regulation of lung vascular P-selectin after systemic activation of complement.

Animals↗

PubMed1997

Mechanisms of acantholysis in pemphigus foliaceus.

Pemphigus foliaceus (PF) is a dermatosis characterized by subcorneal vesicles and pathogenic IgG autoantibodies against desmoglein 1. PF IgG passively transferred into neonatal mice induces a blistering disease that duplicates the key findings of PF. In this study we have used this animal model to investigate the role of complement and IgG valence in triggering blister formation. In the passive transfer experiments, we found that PF IgG, as well as the F(ab')2 and Fab fragments, was capable of inducing the typical subcorneal blistering disease in both complement-deficient and complement-sufficient mice. Moreover, the disease activity in these mice correlated well with the dose of IgG or its proteolytic fragments injected in the animals. We conclude that neither complement activation nor IgG-mediated cell surface antigen crosslinking is required for the induction of acantholysis in the experimental PF model.

Acantholysis↗

PubMed1997

Neutrophil chemotactic activity and C5a following systemic activation of complement in rats.

Using ELISA analysis, rat C5a was stimulated in serum from rats undergoing systemic activation of complement after intravenous infusion of purified cobra venom factor (CVF). Biological (neutrophil chemotactic) activity was also assessed. Serum levels of C5a were directly proportional to the amount of CVF infused. C5a and neutrophil chemotactic activity, peaked by 5 min, then plateaued. In vitro addition of anti-C5a to serum samples of CVF-infused rats totally abolished chemotactic activity, indicating that all biological activity could be ascribed to C5a. Blood neutrophils obtained from CVF-infused animals showed a significant upregulation of CD11b, the increase being reduced (38%) in animals pretreated with anti-C5a. These findings indicate that infusion of CVF into rats produces generation of C5a, all chemotactic activity in serum being related to C5a. The in vivo generation of C5a is, at least inpart, responsible for upregulation of CD11b on blood neutrophils.

Animals↗

PubMed1997

Complement-induced retinal arteriolar occlusions in the cat.

PURPOSE: To develop an animal model of complement-induced retinal vasculopathy and determine whether it resembles Purtscher's retinopathy. METHODS: Intravenous cobra venom factor was used to achieve intravascular activation of the complement system in cats. After a single bolus of cobra venom factor (75 units/kg), retinal blood flow was monitored at regular intervals by fluorescein angioscopy and angiography. RESULTS: Multiple small retinal arteriolar occlusions were present during the initial fluorescein transit of the immediate postinjection fluorescein study in 12 of 12 animals. Small, rapidly moving gaps in the fluorescein column were seen in two thirds of the animals observed continuously by fluorescein angioscopy. Angiographically, the obstructions were transient, and filling of the associated patches of capillary nonperfusion occurred within 3 minutes. Purtscher's-like ischemic retinal infarcts did not develop in any eye. Histopathologic analysis failed to demonstrate the nature of the transient vascular obstructive lesions, but indirect evidence suggested the possibility of granulocyte aggregates. CONCLUSION: Intravascular activation of the complement system produces transient microembolic retinal arteriolar occlusions in the cat. Although this model may represent a mild form of Purtscher's retinopathy, factors in addition to complement activation appear necessary to induce ischemic retinal infarcts.

Angioscopy↗

PubMed1997

Requirement for C5a in lung vascular injury following thermal trauma to rat skin.

Previous studies in rats have shown that deep second degree dermal burns, involving 28-30% of total body surface area, result in systemic complement activation, appearance in plasma of chemotactic activity, sequestration of blood neutrophils in lung capillaries, and development of neutrophil-dependent dermal vascular and lung vascular injury. Although blockade of complement activation or depletion of complement before skin burns has resulted in significant attenuation of tissue injury both locally and distally (in lung), a role for C5a in these events is unclear. In the following studies, we demonstrate the presence of C5a and neutrophil chemotactic activity in serum and in lung homogenates after thermal injury. C5a has also been found in bronchoalveolar lavage fluids of thermally injured animals. Treatment of animals with a polyclonal neutralizing rabbit antibody to rat C5a was lung protective. The protective effects of the antibody (anti-C5a) were associated with diminished vascular permeability changes, as well as reduced tissue build-up of myeloperoxidase. Anti-C5a also prevented up-regulation of lung vascular ICAM-1 (intercellular adhesion molecule-1) in skin-burned rats. These observations indicate that C5a is essential for development of neutrophil accumulation and vascular permeability increases in distant (lung) organs after thermal trauma to skin. The protective effects of anti-C5a in lung, appear to be related to prevention of up-regulation of vascular ICAM-1. Accordingly, C5a may represent a target for clinical approaches in the treatment of organ injury following thermal trauma.

Animals↗

PubMed1997

Partial liquid ventilation protects lung during resuscitation from shock.

Preliminary animal experience with partial liquid ventilation (PLV) suggests that this therapy may diminish neutrophil invasion and capillary leak during acute lung injury. We sought to confirm these findings in a model of shock-induced lung injury. Sixty anesthetized rats were studied. After hemorrhage to an arterial pressure of 25 mmHg for 45 min, animals were resuscitated with blood and saline and treated with gas ventilation alone or with 5 ml/kg of intratracheally administered perflubron. Myeloperoxidase activity was used to measure lung neutrophil content. A permeability index (the bronchoalveolar-to-blood ratio of 125I-labeled albumin activity) quantified alveolar leak. Injury caused an increase in myeloperoxidase that was reversed by PLV (injury = 0.837 +/- 0.452, PLV = 0.257 +/- 0.165; P < 0.01). Capillary permeability also increased with hemorrhage, with a strong trend toward improvement in the PLV group (permeability indexes: injury = 0.094 +/- 0.102, PLV = 0.045 +/- 0.045; 95% confidence interval for injury--PLV: -0.024, 0.1219). We conclude that PLV is associated with a decrease in pulmonary neutrophil accumulation and a trend toward decreased capillary leak after hemorrhagic shock.

Airway Resistance↗

PubMed1996

Requirement and role of C5a in acute lung inflammatory injury in rats.

The complement activation product, C5a, may play a key role in the acute inflammatory response. Polyclonal antibody to rat C5a was used to define the requirements for C5a in neutrophil-dependent inflammatory lung injury after systemic activation of complement by cobra venom factor (CVF) or after intrapulmonary deposition of IgG immune complexes. In the CVF model, intravenous infusion (but not intratracheal instillation) of anti-C5a produced a dose-dependent reduction in lung permeability and in lung content of myeloperoxidase. In C6-deficient rats, CVF infusion caused the same level of lung injury (measured by leak of 125I-albumin) as found in C6-sufficient rats. In the IgG immune complex model of lung injury, anti-C5a administered intratracheally (but not intravenously) reduced in a dose-dependent manner both the increase in lung vascular permeability as well as the buildup of lung myeloperoxidase. Treatment with anti-C5a greatly suppressed upregulation of lung vascular intercellular adhesion molecule-1 (ICAM-1). This was correlated with a substantial drop in levels of TNFalpha in bronchoalveolar fluids. These data demonstrate the requirement for C5a in the two models of injury. In the IgG immune complex model, C5a is required for the full production of TNFalpha and the corresponding upregulation of lung vascular ICAM-1.

Animals↗

PubMed1995

The role of complement in experimental bullous pemphigoid.

Bullous pemphigoid (BP) is a blistering skin disease associated with an IgG autoimmune response directed against the ectodomain of the hemidesmosomal protein, BP180. An animal model of BP has recently been developed by our laboratory based on the passive transfer of rabbit antimurine BP180 antibodies into neonatal BALB/c mice. The experimental animals develop a blistering disease that reproduces all of the key immunopathological features of BP. In the present study we have investigated the role of complement in the pathogenesis of subepidermal blistering in the mouse model of BP. We demonstrate the following. (a) Rabbit anti-murine-BP180 IgG was effective in inducing cutaneous blisters in a C5-sufficient mouse strain, but failed to induce disease in the syngeneic C5-deficient strain; (b) neonatal BALB/c mice, pretreated with cobra venom factor to deplete complement, became resistant to the pathogenic effects of the anti-BP180 IgG; (c) F(ab')2 fragments generated from the anti-BP180 IgG exhibited no pathogenic activity in the mouse model; and (d) histologic evaluation of the skin of mice described in points b and c above showed minimal or no neutrophilic cell infiltration in the upper dermis. Thus, anti-BP180 antibodies trigger subepidermal blistering in this BP model via complement activation. This experimental model of BP should greatly facilitate future studies on the pathophysiology of autoantibody-mediated diseases of the dermal-epidermal junction.

Animals↗

PubMed1994

Phorbol-stimulated influx of extracellular calcium in rat pulmonary artery endothelial cells.

Stimulation of rat pulmonary artery endothelial cells (RPAEC) with phorbol 12-myristate 13-acetate (PMA) resulted in an increase in intracellular calcium ([Ca2+]i). Unlike the response to bradykinin, C5a and tumor necrosis factor-alpha (TNF-alpha) previously reported (15), the PMA-induced increase in [Ca2+]i was predominantly dependent on extracellular calcium. The PMA response paralleled the BAY K 8644-induced, extracellular calcium-dependent increase in [Ca2+]i. Pretreatment of endothelial cells with the protein kinase C inhibitor staurosporine resulted in a concentration-dependent inhibition of the increase in [Ca2+]i in response to PMA. The ability of PMA analogues to induce significant increase in [Ca2+]i paralleled their ability to induce O2- generation in neutrophils. The PMA-induced influx of extracellular Ca2+ was inhibited by the L-channel selective antagonists diltiazem, nifedipine, nicardipine, and verapamil in a dose-dependent manner. Depolarizing conditions induced by high [K+]o enhanced the calcium response to PMA. The data presented are consistent with the hypothesis that PMA-induced increases in [Ca2+]i in endothelial cells are the result of Ca2+ influx through voltage-dependent L-type Ca2+ channels.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1994

Role of selectins in local and remote tissue injury following ischemia and reperfusion.

Ischemia (4-hour) followed by reperfusion (4-hour) of rat hind limbs results in local injury as well as remote (lung) injury. It has recently been shown that injury in this model is neutrophil- and cytokine-dependent and requires the beta 2 integrin adhesion molecules CD11a/CD18 and CD11b/CD18. The role of selectins in events leading to injury (as determined by leakage of albumin and by hemorrhage) was assessed either through the use of blocking antibodies to L-, E- or P-selectins or by the use of oligosaccharides that are reactive with selectins. Lung injury was found to be L- and E-selectin-dependent. When the ischemia and reperfusion times were reduced, lung injury was also found to be P-selectin dependent. In the case of hind limb injury involving the crural muscle mass, injury was L-selectin-dependent but independent of requirements for P- and E-selectin. Injury in both organs was blocked by the infusion of sialylated Lewis pentasaccharide, whereas sialyl-N-acetyllactosamine pentasaccharide failed to protect against injury. In general, when selectin-blocking approaches were protective, there were parallel reductions in tissue content of myeloperoxidase. These data underscore the role of selectins in ischemia-reperfusion injury and suggest that selectin requirements may vary with the vascular bed under study.

Animals↗

PubMed1994

Role of leukocyte adhesion molecules in lung and dermal vascular injury after thermal trauma of skin.

Acute second degree thermal injury of rat skin involving 25 to 30% total body surface of anesthetized rats results at 4 hours in evidence of vascular injury both locally (in skin) and remotely (involving lung). The neutrophil dependency for both types of injury has now been established. Monoclonal antibodies to various adhesion molecules have been used to define the requirements for these molecules in the development of vascular injury. In dermal vascular injury, a requirement for Mac-1 (CD11b/CD18) but not for leukocyte function-associated antigen-1 (LFA-1, CD11a/CD18) has been established. In this model requirements have also been demonstrated for intercellular adhesion molecule-1 (ICAM-1) and E- and L-selectin but not for very late arising antigen-4 (VLA-4) or P-selectin. With respect to lung vascular injury, dual requirements for both leukocyte function-associated antigen-1 and Mac-1 were found as well as for ICAM-1 and E- and L-selectin but not for VLA-4 and P-selectin. In the lung, there was a close correlation between neutrophil content of the tissue (as assessed by myeloperoxidase) and the effects of protective interventions (directed against blocking of adhesion molecules). These data emphasize the roles of beta 2 integrins, selectins (L and E), and ICAM-1 in events that lead to neutrophil-mediated vascular injury of dermis and lung after thermal trauma to skin.

Animals↗

PubMed1994

Acute skin injury releases neutrophil chemoattractants.

BACKGROUND: Progressive or ongoing skin necrosis after traumatic injury is well known. Experimental evidence has associated these events with neutrophil activation and secondary oxidant injury. To determine the mechanism by which neutrophils migrate to a site of injury, cytokine release from injured skin was measured. METHODS: Twenty-five skin biopsy specimens of acute partial thickness skin injuries were compared with uninjured skin of the same patient. Conditioned medium from explanted skin was assayed for tumor necrosis factor (TNF), interleukin-6 (IL-6), and IL-8. RESULTS: Acute skin injury resulted in a significant release of IL-8 but not IL-6 or TNF. In eight patients gradient cytokine release was found; IL-8 levels for partial thickness burn were 26.4 ng/ml, for unburned skin adjacent to the burn were 2.1 ng/ml, and for distal normal skin were 0.2 ng/ml. CONCLUSIONS: IL-8 is released from acutely injured skin; IL-6 and TNF are not. This selective release suggests a mechanism whereby neutrophils are recruited into injured tissue. These neutrophils might then induce further injury, increasing the extent of posttraumatic tissue loss.

Acute Disease↗

PubMed1993

Evidence that NADPH-dependent methemoglobin reductase and administered riboflavin protect tissues from oxidative injury.

NADPH-dependent methemoglobin reductase, first detected in erythrocytes sixty years ago, has subsequently been purified and characterized as a methylene blue reductase and a flavin reductase. The reductase plays no role in methemoglobin reduction under normal conditions, but its activity serves as the basis for the treatment of methemoglobinemia with methylene blue or flavin. On-going studies demonstrate that this cytosolic protein is also present in liver and that its primary structure distinguishes it from other known proteins. The bovine erythrocyte reductase tightly binds hemes, porphyrins, and fatty acids with resulting loss of activity. Pyrroloquinoline quinone serves as a high-affinity substrate of the reductase, suggesting that this naturally-occurring compound may be a physiological substrate. The ability of the reductase to catalyze the intracellular reduction of administered riboflavin to dihydroriboflavin suggested that this system might be exploited to protect tissues from oxidative damage. This hypothesis was supported by our finding that dihydroriboflavin reacts rapidly with Fe(IV)O and Fe(V)O oxidation states of hemeproteins, states that have been implicated in tissue damage associated with ischemia and reperfusion. Preliminary studies demonstrate that, as predicted, administration of low concentrations of riboflavin protects isolated rabbit heart from reoxygenation injury, rat lung from injury resulting from systemic activation of complement, and rat brain from damage caused by four hours of ischemia. Data from these animal studies suggest that flavin therapy holds promise in protecting tissue from the oxidative injuries of myocardial infarction, acute lung injury, stroke, and a number of other clinical conditions.

Animals↗