Skip to main content
NCBI home page
Clinical and Experimental Immunology logoLink to Clinical and Experimental Immunology
. 1998 Mar;111(3):484–490. doi: 10.1046/j.1365-2249.1998.00518.x

A blockade of complement activation prevents rapid intestinal ischaemia-reperfusion injury by modulating mucosal mast cell degranulation in rats

T Kimura 1, A Andoh 1, Y Fujiyama 1, T Saotome 1, T Bamba 1
PMCID: PMC1904891  PMID: 9528887

Abstract

We attempted to define the putative role of complement activation in association with mucosal mast cell (MMC) degranulation in the pathogenesis of rapid intestinal ischaemia-reperfusion (I/R) injury. We prepared complement activity-depleted rats by the administration of the anti-complement agent K-76COOH and the serine-protease inhibitor FUT-175. Autoperfused segments of the jejunum were exposed to 60 min of ischaemia, followed by reperfusion for various time periods, and the epithelial permeability was assessed by the 51Cr-EDTA clearance rate. The number of MMC was immunohistochemically assessed. In control rats, the maximal increase in mucosal permeability was achieved by 30–45 min of reperfusion. This increase was significantly attenuated by the administration of either K-76COONa alone or in combination with FUT-175. In contrast, the administration of carboxypeptidase inhibitor (CPI), which prevents the inactivation of complement-derived anaphylatoxins such as C5a, significantly enhanced the increase in I/R-induced mucosal permeability. These findings were confirmed morphologically by light microscopy and scanning electron microscopy. In addition, the I/R-induced mucosal injury was accompanied by a marked decrease in the number of MMC, and administration of K-76COOH significantly inhibited this change. These results indicate that complement activation and the generation of complement-derived anaphylatoxins are key events in I/R-induced mucosal injury. It is likely that intestinal I/R-induced mucosal injury may be partially mediated by MMC activation associated with the complement activation.

Keywords: complement, mucosal mast cell, anaphylatoxin

INTRODUCTION

Ischaemia-reperfusion (I/R) of the small bowel is associated with both increased microvascular permeability and mucosal barrier dysfunction, and results in systemic shock under clinical and experimental conditions. There is an increasing number of studies indicating that the mucosal and vascular dysfunction induced by hypoxic-reoxygenation damage is associated with the release of toxic factors into the systemic circulation, leading to acute circulatory collapse [1–4]. However, the mechanisms mediating this mucosal barrier dysfunction after reperfusion are not fully understood.

The complement system has evolved into a major humoral immune defence mechanism, but its participation in a variety of inflammatory responses is not entirely beneficial. Indeed, several experimental reports have suggested that complement activation may be involved in I/R injury in the heart [5,6], lung [7], skeletal muscle [8] and intestine [9]. One possible mechanism for complement-mediated tissue injury depends on the generation of a set of biologically active peptides, the anaphylatoxins (C3a, C4a and C5a). Of these peptides, C5a is the major factor and the most potent in biological activity. The generation of C5a plays a central role in the systemic and local inflammation associated with complement activation. In addition, C5a acts as a potent stimulator for mast cells. This peptide binds to receptors on the surface of mast cells, and initiates the release of vasoactive amines from intracellular granules [10]. The histamine and serotonin released then act on the smooth muscle to cause constriction, and on the small vessels to increase vascular permeability. Thus, activation of the complement cascade induces mast cell degranulation and enhances tissue injury.

The small intestine contains mucosal mast cells (MMC) at a high density and at close proximity to the microvasculature of the villus [11]. These MMC are the local source of large amounts of vasoactive mediators postulated to play important functions in I/R-induced tissue injury. The purpose of the present study was to elucidate the role of the interaction between complement activation and MMC activation in the formation of rapid mucosal barrier dysfunction after intestinal I/R. This was mainly tested by determining the action of anti-complementary agent K-76COONa and serine-protease inhibitor FUT-175. K-76COONa has been established as a potent inhibitor of complement activation at the C5 step, and blocks the generation of C5a [12–14]. FUT-175 has been reported to be a potent serine protease inhibitor, and exerts various anti-inflammatory effects, including inhibition of the activities of complement components (C1r, C1s, C3 and factor B) [15–18]. Thus, the i.p. administration of K-76COONa and FUT-175 potently attenuates the activity of both classical and alternative complement activation pathways [19].

MATERIALS AND METHODS

Surgical preparation

These experiments were performed on male Wistar rats (200–350 g) anaesthetized with sodium pentobarbital (Nakalai, Kyoto, Japan). The superior mesenteric artery was isolated, and 12 to 15-cm loop of jejunum was externalized; the blood vessels remained intact. The jejunal loop was fitted with inflow and outflow tubes to allow the perfusion of warm Tyroid solution at a rate of 0.4 ml/min. Jejunal ischaemia was subsequently induced by clamping the superior mesenteric artery for 60 min, after which the clamp was removed, and then reperfusion was monitored for up to 120 min. Sham-operated rats were treated in an identical fashion with the omission of vascular occlusion.

Measurement of epithelial permeability

51Cr-EDTA was injected via the jugular vein and allowed to equilibrate. The luminal perfusate was collected during both ischaemia and reperfusion. Plasma samples were also collected. The plasma-to-lumen clearance of 51Cr-EDTA was calculated, as described by Kanwar & Kubes [9]:

graphic file with name cei0111-0484-m1.jpg

where the clearance of 51Cr-EDTA is expressed as ml/min per 100 g, ct/minp is the counts/min per ml of the luminal perfusate, Pr is the perfusion rate (ml/min), ct/minpl is the counts/min per ml of plasma, and wt is the weight of the intestinal segment (g).

Histological techniques

The specimens were immediately fixed in 10% formaldehyde-saline solution, followed by sectioning and haematoxylin–eosin staining. For the immunohistochemical detection of MMC, samples were reacted with a polyclonal rabbit anti-rat mast cell protease II (RMCP II; Moredun, Edinburgh, UK) [20] antibody, followed by reaction with the immunoperoxidase streptavidin-biotin complex system (Dako, Glostrup, Denmark). The number of MMC per 100 villi per animal were counted by microscopic examination for the evaluation of the degranulation of MMC.

Scanning electron microscopy

The samples were fixed in 2.5% glutaraldehyde in 0.1 m sodium cacodylate buffer pH 7.4 for 4 h, followed by post-fixation in 0.1 m OsO4 pH 7.4 for 1 h. The samples were dehydrated in an ascending ethanol series, and critical point-dried by the CO2 method. After drying, the samples were ion-sputtered to minimize charging with a thin platinum coating, using a single-gun, argon ion beam sputterer operated at 4 mA at an accelerating voltage of 10 keV for 6 min. Scanning electron microscopy (SEM) was then performed at 1.5 keV with a Hitachi S-570 (Tokyo, Japan) electron microscope.

Carboxypeptidase inhibitor

DL-2-mercaptomethyl-3-guanidinoethyl thiopropanoic acid (Calbiochem, La Jolla, CA) is a carboxypeptidase inhibitor which prevents the inactivation of C5a and C3a. The carboxypeptidase inhibitor was administered intraperitoneally at the start of ischaemia, at a concentration of 100 mg/kg, as previously described [21].

Anti-complement agents

The sodium salt of K-76 monocarboxylic acid (K-76COOH) and FUT-175 were obtained from Otsuka Pharmaceutical Co. Ltd. (Tokushima, Japan) and Torii Co. Ltd. (Tokyo, Japan), respectively. To block complement activity, each rat was injected intraperitoneally with K-76COONa (200 mg/kg) or a combination of K-76COONa and FUT-175 (40 mg/kg) at the start of ischaemia, as previously described [19].

Myeloperoxidase assay

The intestinal neutrophil content was assessed by a calorimetric assay of the neutrophil enzyme, myeloperoxidase (MPO), by the standard method as previously described [22,23].

RESULTS

The mucosal permeability, as determined by 51Cr-EDTA clearance, was assessed for 120 min (Fig. 1). Treatment with 60 min of ischaemia alone did not induce any significant changes. Within 15 min of reperfusion, however, there was a rapid and significant increase in mucosal permeability. This change reached a maximum as early as 30–45 min after the start of reperfusion, and was sustained for ≈ 120 min. The administration of K-76COONa (200 mg/kg) alone or in combination with FUT-175 (40 mg/kg) significantly reduced this I/R-induced increase in mucosal permeability induced by 30–60 min of reperfusion (Fig. 1). In particular, the combination of K-76COONa plus FUT-175 demonstrated a dramatic inhibitory effect which was significantly stronger than K-76COOH alone at 45 min of reperfusion (P < 0.05). This indicates that complement activation participates in the pathogenesis of the rapid mucosal injury induced by I/R treatment. To investigate further the role of complement activation, we tested the effects of carboxypeptidase inhibitor (CPI). CPI has been established as a potent inhibitor of the degradation of complement-derived anaphylatoxins [21]. As shown in Fig. 2, the administration of CPI significantly enhanced the increase in mucosal permeability induced by the I/R treatment (P < 0.05), suggesting an important role for anaphylatoxin generation in I/R-induced mucosal injury.

Fig. 1.

Fig. 1

Open in a new tab

Changes in epithelial permeability induced by ischaemia-reperfusion (I/R) treatment in rats. Open and closed bars indicate animals treated with sham operation and I/R, respectively. Hatched and cross-hatched bars indicate animals treated with K-76COONa alone (200 mg/kg) and combination of K-76COONa (200 mg/kg) and FUT-175 (40 mg/kg) before I/R, respectively. Data expressed as mean ± s.d.; n = 5 per group. *P < 0.01 versus respective sham operation value; **P < 0.01versus respective I/R value.

Fig. 2.

Fig. 2

Open in a new tab

Effects of carboxypeptidase inhibitor (CPI) on epithelial permeability induced by ischaemia-reperfusion (I/R) treatment. ▪, Animals treated with I/R; □, animals treated with CPI. Data expressed as mean ± s.d.; n = 5 per group. *P< 0.01 versus respective I/R treated value.

The mucosal damage induced by 30 min of reperfusion was assessed histologically by light microscope and SEM. As demonstrated in Fig. 3, I/R treatment induced severe mucosal destruction, as represented by disruption of the villus tip, loss of villus height, dilated capillaries with haemorrhage, and ulceration. The severity of these changes was potently enhanced by the administration of CPI, following which marked disintegration of villi was observed. On the other hand, the administration of K-76COONa had a remarkable protective effect; there was a residual subepithelial space at the villus tip with moderate lifting of the epithelium. SEM also detected similar phenomena (Fig. 4). The smooth and regular structures of villi in the controls were destroyed by I/R treatment, and the disruption of the villus tip and overflow of erythrocytes were clearly detected. Treatment with K-76COONa alone completely abrogated these changes, and there was no detectable morphological damage on SEM observation (Fig. 4D).

Fig. 3.

Fig. 3

Open in a new tab

Light micrographs of H–E-stained sections of rat intestinal mucosa after ischaemia-reperfusion (I/R) treatment. (A) Sham. (B) I/R-treated. (C) Carboxypeptidase inhibitor (CPI) treated. (D) K-76COONa-treated. (Mag. × 100.)

Fig. 4.

Fig. 4

Open in a new tab

Scanning electron micrographs of mucosal villi after ischaemia-reperfusion (I/R) treatment. (A) Sham. (B,C) I/R-treated. (D) K-76COONa-treated. (Mag. A,B,D × 400; C, × 2000.)

To make clear the relationship between complement activation and I/R-induced mucosal injury, we focused on the role of anaphylatoxin-induced MMC activation. As demonstrated in Fig. 5, immunoreactive MMC were stained with anti-RMCP II antibody. This showed that I/R treatment remarkably decreased the number of MMC, whereas K-76COOH completely attenuated this response. In Fig. 6, the number of MMC/100 villi was counted under light microscope. The number of MMC at 30 min of reperfusion was significantly decreased by I/R treatment compared with controls (Fig. 6a; P < 0.01), but the administration of K-76COOH either alone or in combination with FUT-175 clearly prevented this alteration (Fig. 6a). These results indicate that the decrease in MMC number was closely linked to complement activation. To evaluate the effects of the infiltrating neutrophils, we assessed the mucosal MPO activity (Fig. 6b). Contrary to the above results, we could not detect any significant differences in MPO activity. We therefore conclude that neutrophil infiltration has little or no effect on the pathogenesis of rapid I/R-induced mucosal injury.

Fig. 5.

Fig. 5

Open in a new tab

Immunoreactive mucosal mast cells (MMC) in the intestinal mucosa. MMC were clearly detected by anti-RMCP II antibody. (A) Sham. (B) Ischaemia-reperfusion (I/R)-treated. (C) K-76COONa-treated.

Fig. 6.

Fig. 6

Open in a new tab

Changes in the number of mucosal mast cells (MMC) (a) and myeloperoxidase (MPO) activity (b). Number of resting MMC per 100 villi was counted under light microscope (n = 5 per group), and MPO activity was assessed by standard method (n = 5 per group). **P< 0.01; *P< 0.05.

DISCUSSION

The objective of this study was to investigate the potential role of complement activation and the subsequent MMC activation in the pathogenesis of rapid (30–45 min of reperfusion) intestinal I/R injury. Our approach was to compare the response of control animals against complement-depleted animals, which had a residual serum complement activity of the alternative pathway of < 5% of control values [19].

In the present study we used the anti-complement agent K-76COONa and serine-protease inhibitor FUT-175 to define the role of complement activation in intestinal I/R injury. Previous studies have shown that the administration of these agents in vivo causes a potent reduction in the haemolytic activity of both classical and alternative pathways; these agents induced an almost complete inhibition of the alternative pathway, and reduced classical pathway activity by 50–60% [19]. In our experiments, the administration of K-76COOH alone markedly prevented the increase in mucosal permeability induced by I/R treatment, indicating an important role of complement activation in I/R-induced mucosal injury. Furthermore, the mucosal permeability at 45 min of reperfusion indicated that the effects of combining K-76COONa with FUT-175 were significantly stronger than K-76COONa alone. While various anti-inflammatory effects of FUT-175 have been reported, it is suggested that the potent inhibitory effect of FUT-175 on complement activation may participate in these responses. Judging by the mode of action of K-76COONa and FUT-175 on complement activation, alternative pathway activation might play a more important role in I/R-induced mucosal injury. This finding is compatible with the previous study reporting that complement activation during the early reperfusion phase may be caused by reactive oxygen species via the alternative pathway [24].

Next, we tested the effects of CPI, which prevents the degradation of complement-derived anaphylatoxins. It is well known that generated anaphylatoxins are rapidly inactivated by carboxypeptidases in plasma [25], and as shown in a previous study the administration of CPI (100 mg/kg) completely inhibits plasma carboxypeptidase activity in vivo [21]. Indeed, the administration of CPI enhanced I/R-induced mucosal injury, as evaluated by the increase in permeability and the progress of histological damage. Thus, the results from both the anti-complement agents and CPI treatment indicate that the increase in I/R-induced mucosal injury is closely associated with complement activation and the generation of anaphylatoxins. The importance of complement activation in intestinal I/R injury has also been demonstrated by a previous report using a long period of reperfusion (180 min) [7]. However, we emphasize here that complement activation modulates the generation of I/R-induced mucosal injury more rapidly (30 min of reperfusion). There are many different inflammatory responses, such as neutrophil infiltration and chemical mediator release, between early (30 min) and late (180 min) phase reperfusion: thus the responses observed at the late phase may be more complicated than at the early phase. Our data therefore indicate more clearly that complement activation is critical in the injury process following I/R. However, the depletion of complement activation reduced, but did not completely prevent, the increase in mucosal permeability. It seems likely that a variety of chemical mediators and signals besides complement activation are involved in the pathogenesis of I/R injury, and these factors can compensate for the absence of one mediator.

I/R treatment induced a marked decrease in the number of MMC in the intestinal mucosa, suggesting that I/R treatment enhanced the degranulation of MMC. Since activated mast cells can release preformed chemical mediators, including histamine [1,2], leukotrienes [4] and platelet-activating factor (PAF) [3] within 15 s after stimulation [10], it is possible that part of the tissue injury observed in our model might be mediated by these factors released from the activated MMC. Although it has previously been reported that neutrophils also mediate the generation of I/R injury [9], we could not detect this in our model. There were no differences in MPO activity in our experiments, and these results were compatible with several reports of hepatic I/R injury [26,27]. In these reports, early neutrophil infiltration had no impact on hepatic I/R injury because the cells were not fully activated yet [26,27]. However, after about 5–6 h of reperfusion, they can contribute to the post-ischaemic oxidant stress and injury. In our model, it is likely that the 30 min of reperfusion was too short to increase the infiltration and/or activation of neutrophils [28,29]. Although our data suggest a major role for MMC but not neutrophils in the pathogenesis of intestinal I/R injury, the question remains which mediator is responsible for the stimulation of the MMC. Several possibilities can account for the increased degranulation of MMC induced by I/R treatment. In the present study, we focused on the role of complement activation and the generation of anaphylatoxin. It is well known that complement-derived anaphylatoxins (C5a, C3a and C4a) are potent proinflammatory mediators. In addition, anaphylatoxins act as potent activators of mast cells and can induce degranulation [10,30]. In this study, since the depletion of complement activity significantly attenuated the degranulation of MMC, it should be noted that MMC activation is closely associated with the rapid generation of complement-derived anaphylatoxins.

In conclusion, the I/R experiments with complement-depleted animals and the effects of CPI strongly suggest the involvement of complement activation in the generation of I/R-induced mucosal injury. In particular, it seems likely that the degranulation of MMC associated with complement activation is an important process for the induction of mucosal damage. Thus, based on our data, we propose the following mechanism for the contribution of complement activation and MMC to the pathogenesis of rapid intestinal I/R injury. Intestinal I/R treatment causes an initial activation of the complement cascade mainly via the alternative pathway by reactive oxygen species and generates anaphylatoxins such as C5a. These anaphylatoxins subsequently induce the activation of resident MMC, resulting in the release of chemical mediators which causes the mucosal injury. This initial injury leads to more complement activation, with further enhancement of the injury in a self-aggravating process. In addition, these activated complement factors may contribute directly or indirectly to subsequent MMC activation and further tissue injury. Our data indicate that in the pathogenesis of intestinal rapid I/R injury, complement activation and MMC activation are closely linked. It is suggested that a blockade of the complement cascade may be important for the development of therapeutic strategies.

References

  • 1.Boros M, Kaszaki J, Nagy S. Histamine release during intestinal ischemia-reperfusion: role of iron ions and hydrogen peroxide. Circ Shock. 1991;35:174–80. [PubMed] [Google Scholar]
  • 2.Boros M, Kaszaki J, Nagy S. Oxygen free radical induced histamine release during intestinal ischemia and reperfusion. Eur Surg Res. 1989;21:297–304. doi: 10.1159/000129042. [DOI] [PubMed] [Google Scholar]
  • 3.Kubes P, Ibbotson G, Russel J, et al. Role of platelet-activating factor in ischemia/reperfusion-induced leukocyte adherence. Am J Physiol. 1990;259:G300–G5. doi: 10.1152/ajpgi.1990.259.2.G300. [DOI] [PubMed] [Google Scholar]
  • 4.Lehr HA, Guhlmann A, Nolte D, et al. Leukotrienes as mediators in ischemia-reperfusion injury in a microcirculation model in the hamster. J Clin Invest. 1991;87:2036–41. doi: 10.1172/JCI115233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Smith Iiief, Griswold DE, Egan JW, et al. Reduction of myocardial reperfusion injury with human soluble complement receptor type 1 (BRL 55730) Eur J Pharmacol. 1993;236:477–81. doi: 10.1016/0014-2999(93)90487-3. [DOI] [PubMed] [Google Scholar]
  • 6.Ito BR, Engler RL, Del Balzo U. Role of cardiac mast cells in complement C5a-induced myocardial ischemia. Am J Physiol. 1993;264:H1346–H54. doi: 10.1152/ajpheart.1993.264.5.H1346. [DOI] [PubMed] [Google Scholar]
  • 7.Hill J, Lindsay TF, Ortiz F, et al. Soluble complement receptor type I ameliorates the local and remote organ injury after intestinal ischemia-reperfusion in the rat. J Immunol. 1992;149:1723–8. [PubMed] [Google Scholar]
  • 8.Pemberton M, Anderson G, Vetvicka V, et al. Microvascular effects of complement blockade with soluble recombinant CR1 on ischemia/reperfusion injury of skeletal muscle. J Immunol. 1993;150:5104–13. [PubMed] [Google Scholar]
  • 9.Kanwar S, Kubes P. Mast cells contribute to ischemia-reperfusion-induced granulocyte infiltration and intestinal dysfunction. Am J Physiol. 1994;267:G316–G21. doi: 10.1152/ajpgi.1994.267.2.G316. [DOI] [PubMed] [Google Scholar]
  • 10.El-Lati SG, Dahinden CA, Church MK. Complement peptides C3a- and C5a-induced mediator release from dissociated human skin mast cells. J Invest Dermatol. 1994;102:803–6. doi: 10.1111/1523-1747.ep12378589. [DOI] [PubMed] [Google Scholar]
  • 11.Bacci S, Faussone-Pellegrini MS, Mayer B, et al. Distribution of mast cells in human ileocecal region. Dig Dis Sci. 1995;40:357–65. doi: 10.1007/BF02065422. [DOI] [PubMed] [Google Scholar]
  • 12.Hong K, Kinoshita T, Miyazaki W, et al. An anticomplementary agent, K-76 monocarboxylic acid: its site and mechanism of inhibition of the complement activation cascade. J Immunol. 1979;122:2418–23. [PubMed] [Google Scholar]
  • 13.Hong K, Kinoshita T, Kitajima H, et al. Inhibitory effect of K-76 monocarboxylic acid, an anticomplementary agent, on the C3b inactivator system. J Immunol. 1981;127:104–8. [PubMed] [Google Scholar]
  • 14.Hong K, Kinoshita T, Inoue K. Simple methods for preparing EAC1,4b,2a,3b and EAC4b,3b with human or guinea pig complement components using an anticomplementary agent, K-76 monocarboxylic acid. J Immunol. 1981;127:109–14. [PubMed] [Google Scholar]
  • 15.Ikari N, Sakai Y, Hitomi Y, et al. New synthetic inhibitor to the alternative complement pathway. Immunol. 1983;49:685–91. [PMC free article] [PubMed] [Google Scholar]
  • 16.Fujii S, Hitomi Y. New synthetic inhibitors of C1r, C1 esterase, thrombin plasmin kallikrein and trypsin. Biochim Biophys Acta. 1981;661:342–5. doi: 10.1016/0005-2744(81)90023-1. [DOI] [PubMed] [Google Scholar]
  • 17.Hitomi Y, Fujii S. Inhibition of various immunological reactions in vivo by a new synthetic complement inhibitor. Int Archs Allergy Appl Immunol. 1982;69:262–7. doi: 10.1159/000233181. [DOI] [PubMed] [Google Scholar]
  • 18.Ikari N, Hitomi Y, Niinobe M, et al. Studies on esterolytic activity of alternative complement component factor B. Biochim Biophys Acta. 1983;742:318–23. doi: 10.1016/0167-4838(83)90317-5. [DOI] [PubMed] [Google Scholar]
  • 19.Miyagawa S, Shirakura R, Matsumiya G, et al. Prolonging discordant xenograft survival with anticomplement reagents K76COOH and FUT-175. Transplantation. 1993;55:709–13. doi: 10.1097/00007890-199304000-00004. [DOI] [PubMed] [Google Scholar]
  • 20.Kido H, Fukusen N, Katunuma N. Chymotrypsin- and trypsin-type serine proteases in rat mast cells: properties and functions. Arch Biochem Biophys. 1985;239:436–43. doi: 10.1016/0003-9861(85)90709-x. [DOI] [PubMed] [Google Scholar]
  • 21.Ikai M, Itoh M, Joh T, et al. Complement plays an essential role in shock following intestinal ischaemia in rats. Clin Exp Immunol. 1996;106:156–9. doi: 10.1046/j.1365-2249.1996.d01-817.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Kanwar S, Wallace JL, Befus D, et al. Nitric oxide synthesis inhibition increases epithelial permeability via mast cells. Am J Physiol. 1994;266:G222–9. doi: 10.1152/ajpgi.1994.266.2.G222. [DOI] [PubMed] [Google Scholar]
  • 23.Kubes P, Grisham MB, Barrowman JA, et al. Leukocyte-induced vascular protein leakage in cat mesentery. Am J Physiol. 1991;261:H1872–9. doi: 10.1152/ajpheart.1991.261.6.H1872. [DOI] [PubMed] [Google Scholar]
  • 24.Shingu M, Nonaka S, Nishimukai H, et al. Activation of complement in normal serum by hydrogen peroxide and hydrogen peroxide-related oxygen radicals produced by activated neutrophils. Clin Exp Immunol. 1992;90:72–78. doi: 10.1111/j.1365-2249.1992.tb05834.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25.Plummer TH, Ryan TJ. A potent mercapto bi-product analogue inhibitor for human carboxypeptidase N. Biochem Bioph Res Commun. 1981;98:448–54. doi: 10.1016/0006-291x(81)90860-3. [DOI] [PubMed] [Google Scholar]
  • 26.Jaeschke H, Farhood A. Neutrophil and Kupffer cell-induced oxidant stress and ischemia-reperfusion injury in rat liver. Am J Physiol. 1991;260:G355–62. doi: 10.1152/ajpgi.1991.260.3.G355. [DOI] [PubMed] [Google Scholar]
  • 27.Jaeschke H, Bautista AP, Spolarics Z, et al. Superoxide generation by Kupffer cells and priming of neutrophils during reperfusion after hepatic ischemia. Free Radical Res Comms. 1991;15:277–84. doi: 10.3109/10715769109105223. [DOI] [PubMed] [Google Scholar]
  • 28.Jaeschke H, Bautista AP, Spolarics Z, et al. Superoxide generation by neutrophils and Kupffer cells during in vivo reperfusion after hepatic ischemia in rats. J Leukocyte Biol. 1992;52:377–82. doi: 10.1002/jlb.52.4.377. [DOI] [PubMed] [Google Scholar]
  • 29.Jaeschke H, Farhood A, Smith CW. Neutrophils contribute to ischemia/reperfusion injury in rat liver in vivo. FASEB J. 1990;4:3355–9. [PubMed] [Google Scholar]
  • 30.Fukuoka Y, Hugli TE. Anaphylatoxin binding and degradation by rat peritoneal mast cells: mechanisms of degranulation and control. J Immunol. 1990;145:1851–8. [PubMed] [Google Scholar]

Articles from Clinical and Experimental Immunology are provided here courtesy of British Society for Immunology

RESOURCES