Tumor necrosis factor induced acute lung leak in rats: Less than with interleukin-1

Although local tumor necrosis factor- (TNF) release by alveolar macrophages has been postulated to contribute to the development of acute respiratory distress syndrome (ARDS), the effects of instilling TNF intratracheally on the development of acute lung leak are not known. Our goal was to determine the effect of intratracheally administered TNF on the development of acute lung leak in rats. We found that rats given TNF (500 ng) 5 hours previously intratracheally had increased (p<0.05) lung lavage cytokine induced neutrophil chemoattractant (CINC) concentrations, lung lavage neutrophils, lung myeloperoxidase (MPO) activity, and lung leak compared to saline-treated control rats. However, all of the responses following TNF instillation were much lower than the responses to interleukin-1 (IL-1) instillation. For example, instilling 50 ng of IL-1 caused 6.4 times the increases in lung lavage CINC concentrations, 15.5 times the increase in lung lavage neutrophils, 3.6 times the increase in lung MPO activity and 3.8 times the increase in lung leak caused by giving 500 ng of TNF intratracheally. Cotreatment with TNF-binding protein decreased both lung MPO and lung leak increases in rats given TNF intratracheally. These observations suggest that locally elevated levels of TNF may induce lung neutrophil recruitment and acute lung leak but that IL-1 is a much more potent agent than TNF in causing lung neutrophil accumulation and lung leak.Parker B. Francis Fellow in Pulmonary Research.     

Rat CINC, a member of the interleukin-8 family, is a neutrophil-specific chemoattractant in vivo

Rat cytokine-induced neutrophil chemoattractant (CINC) is a member of the IL-8 family and its human counterpart is MGSA/gro. Rat neutrophil responses in vitro to rat CINC, human IL-8, and human MGSA/gro were studied. CINC concentrations as low as 1 nM induced apparent chemotaxis of rat neutrophils, but human IL-8 and MGSA/gro required concentrations one or two orders higher than that of CINC to attract neutrophils. These data indicate that human IL-8 and MGSA/gro cannot sufficiently substitute for rat counterparts such as CINC in rats. Therefore, the effect of rat CINC on rats was studied. Intradermally injected 10(-10)-10(-7) M CINC dose-dependently caused infiltration of neutrophils. Significant migration of neutrophils appeared by 30 min, and maximum infiltration was observed around 1-2 hr after the injection. CINC induced quick and transient neutrophil accumulation without lymphocyte and monocyte migration or edema formation. CINC, a member of the IL-8 family but a counterpart of human MGSA/gro-related proteins, is a specific neutrophil chemoattractant and can be distinguished from IL-8, which is a chemotactic factor for lymphocytes and neutrophils.     

Interleukin-1β-induced neutrophil recruitment and acute lung injury in hamsters

Administering recombinant interleukin-1 (IL-1) intratracheally caused lung neutrophil accumulation and lung injury in hamsters. The percentage of leukocytes that were neutrophils increased progressively in lavages from lungs of hamsters given 25, 50, or 100 ng IL-1 intratracheally 2 h before. Lung injury, reflected by increased lung lavage protein concentrations and lung lavage hemoglobin concentrations, increased 2 h after administering 100 ng IL-1. Lung injury, reflected by lung wet weight/body weight ratios, followed similar patterns, with significant increases occurring 2 h after insufflating 50 or 100 ng IL-1. Our results indicate that increased concentrations of IL-1 in lung airways can cause neutrophil recruitment and lung injury in hamsters. This mechanism may contribute to the development of lung neutrophil accumulation and lung injury that characterizes ARDS patients who have increased airway levels of IL-1.     

Alveolar type II cell abnormalities and peroxide formation in lungs of rats given IL-1 intratracheally

Acute lung injury (ALI) is characterized by increased lung levels of proinflammatory cytokines, inflammation, oxidative stress, edema, and impaired gas exchange. Notably, ALI patients also exhibit pulmonary surfactant abnormalities, including increased levels of phospholipids in their lung lavages. In the present study, to assess early alterations of the lung surfactant system in ALI, we induced inflammation and acute lung injury in rats by administering interleukin-1 (IL-1) intratracheally. Five h after IL-1 instillation, we examined lung tissue ultrastructure by electron microscopy using both routine staining methods and cerium chloride staining to localize hydrogen peroxide (H₂O₂) histologically. We also measured lung lavage phospholipid levels, lung tissue -glutamyl transpeptidase (GGT) activities (a marker of oxidative stress), and arterial blood oxygen tensions. We observed that lungs of rats given IL-1 intratracheally had increased neutrophil accumulation, increased H₂O₂ production, and increased alveolar type II (ATII) pneumocyte ultrastructural abnormalities compared to rats given saline intratracheally. Intratracheal instillation of IL-1 also increased phospholipid levels in the bronchoalveolar lavage (BAL), possibly as a consequence of the abnormal discharge of lamellar bodies into the alveolar lumen. In addition, IL-1-insufflated rats had increased lung GGT levels and impaired blood oxygenation compared to saline-insufflated rats. Treatment with mepacrine decreased lung neutrophil accumulation, ultrastructural lung abnormalities, lung lavage phospholipid levels, lung tissue GGT levels, and blood oxygenation impairment in rats given IL-1 intratracheally, suggesting a possible relationship between these events. Our results indicate that IL-1-induced acute lung injury in rats is marked by neutrophil-dependent oxidative stress, ATII cell defects, abnormal discharge of lamellar body phospholipids, and impaired blood oxygenation.     

Aging Reduces the Expression of Lung CINC and MCP-1 mRNA in a P. aeruginosa Rat Model of Infection

We investigated dynamic changes of inflammatory cell infiltration and expression of cytokine-induced neutrophil chemoattractant (CINC) and monocyte chemoattractant protein-1 (MCP-1) mRNA in aged rats with Pseudomonas aeruginosa pulmonary infection. Disease manifestation and lung tissue pathology (lesion dispersion, inflammatory reactions, tissue edema and bleeding) were more severe in aged rats than young rats. At various time points, lung tissue polymorphonuclear neutrophil and mononuclear macrophage numbers were lower in the aged group than the young group (P < 0.05), and at 24 h there was no difference in mononuclear macrophage numbers. After inoculation with P. aeruginosa, CINC and MCP-1 mRNA expression increased in both groups, but the peak lagged in old rats compared with young. Thus, aging can reduce the expression of CINC and MCP-1 mRNA in lung tissues, and reduce the infiltration of neutrophils and monocyte–macrophages induced by CINC and MCP-1. This might lead to increased risk of pneumonia in elderly patients.     

Appearance of cytokine-induced neutrophil chemoattractant isoforms and immunolocalization of them in lipopolysaccharide-induced acute lung inflammation in rats

OBJECTIVE AND DESIGN: Recently, rat cytokine-induced neutrophil chemoattractant (CINC), which belongs to the interleukin-8 family, was grouped into four isoforms, CINC-1, CINC-2a, CINC-2beta, and CINC-3. To determine the major component and the source of CINC in airways, we investigated the change in appearance of CINC isoforms after exposure of rats to lipopolysaccharide. METHODS: Male Sprague-Dawley rats, 8-10 weeks old, were used in the present study. Bronchoalveolar lavage (BAL) was performed at 1, 2, 4, 6, 12, and 24 h after lipopolysaccharide inhalation (4 mg/ml for 30 min). The concentrations of each specific rat CINC in the BAL supernatant were measured by use of commercially available kits. Furthermore, lung tissue was employed for immunohistochemical staining of CINCs (CINC-1, -2alpha,-2beta, and -3) using the streptavidin-biotin technique. RESULTS: Inhalation of lipopolysaccharide caused increases in CINC-1, CINC-2aalpha, and CINC-3 in BAL fluids, whereas CINC-2beta was not detected. The increases in CINC-2a and CINC-3 were less than the increase in CINC-1. Positive immunohistochemical staining for CINC-1 was detected in bronchial noncilliated cells and in certain neutrophils that had infiltrated into the submucosa. CONCLUSIONS: These findings suggest that CINC-1 is the major isoform among the four CINCs in lipopolysaccharide-induced acute lung inflammation in rats. Its sources are likely to be bronchial noncilliated cells and certain infiltrating neutrophils.     

Cytokine-induced neutrophil chemoattractant production by primary rat alveolar type II cells

This study was designed to determine the production of the chemokine cytokine-induced neutrophil chemoattractant (CINC) by primary rat alveolar type II (ATII) cells upon stimulation with exogenous and endogenous proinflammatory factors. Cultures of primary rat ATII cells were exposed to lipopolysaccharide (LPS), interleukin-1 beta (IL-1) or tumor necrosis factor-alpha (TNF) over a 16 hour period and the production of CINC both apically and basolaterally was measured by ELISA. Compared to unstimulated (UNS) cultures, LPS, IL-1 and TNF were found to significantly increase the level of CINC detected in culture by two, four and sixteen hours post stimulation, respectively. ATII cells also demonstrated a polar secretion of CINC. The accumulation of CINC basolaterally was significantly more than apically; 133%, 45%, 117% and 123% for UNS, IL-1, LPS and TNF respectively. We demonstrated that primary rat ATII cells may participate in the chemokine network during inflammation by the production of CINC upon stimulation with endogenous and exogenous factors.     

Effect of rat CINC/gro, a member of the interleukin-8 family, on leukocytes in microcirculation of the rat mesentery.

Rat cytokine-induced neutrophil chemoattractant (CINC) is a member of the IL-8 family, and its human counterpart is gro/MGSA but not IL-8. We ascertained that chemically synthesized CINC was comparable to native CINC/gro with regard to chemotactic activity for rat neutrophils and studied the effect of synthesized CINC/gro on circulating leukocytes in microvascular vessels of rat mesentery. Exposure of rat mesentery to 10(-8)M authentic CINC/gro induced neutrophil adherence to and extravasation from postcapillary venules (PCVs) but not from capillaries or arterioles. CINC/gro concentrations as low as 10(-10) M were effective in causing neutrophil adherence. Neutrophils adhered to thin PCVs (mean diameter, approximately 25 microns) after exposure to CINC/gro for 15 min. The mean diameters of the PCV with adherence of neutrophils after exposure to CINC/gro for 30 and 60 min were 37 and 43 microns, respectively. The diameters of PCV with extravasation of neutrophils also increased in a time-dependent manner. The starting position of adherence of neutrophils was approximately 25-50 microns away from the upper junction of two vessels and remained virtually unchanged during exposure to CINC/gro for 60 min. However, the distance from the start to the end of neutrophil adherence increased in a time-dependent manner. The effect of CINC/gro on adherence and extravasation of leukocytes was neutrophil specific since other leukocytes such as lymphocytes and monocytes were not identified among the adherent and extravasated leukocytes.     

Level of neutrophil chemotactic factor CINC/gro, a member of the interleukin-8 family, associated with lipopolysaccharide-induced inflammation in rats.

Rat cytokine-induced neutrophil chemoattractant (CINC), which is a counterpart of human gro and belongs to the interleukin-8 family, has been quantified by a new sandwich enzyme-linked immunosorbent assay. Administration of lipopolysaccharide (LPS) into an air pouch performed by subcutaneous injection of air caused inflammation and severe neutrophil infiltration. After the LPS injection, changes in the concentration of CINC/gro, chemotactic activity, and the number of neutrophils in the air pouch exudate were determined. The chemotactic activity of neutrophils was augmented before practical neutrophil infiltration. More than half of the chemotactic activity was neutralized by the antisera. The time kinetics of the level of CINC/gro coincided with the changes in chemotactic activity. The maximal level of rat CINC/gro was 85 ng/ml, which is sufficient to cause neutrophil migration in vitro and in vivo as described previously. These data suggest that rat CINC/gro is a functional chemoattractant for neutrophils in LPS-induced inflammation in rats.     

Stimulatory effect of the herbal medicine Keishi-bukuryo-gan on a cytokine-induced neutrophil chemoattractant, in rat ovarian cell culture

PROBLEM AND METHOD OF STUDY: We investigated the effects of Keishi-bukuryo-gan, a Japanese herbal medicine, and its crude ingredients in relation to the production of cytokine-induced neutrophil chemoattractant (CINC/gro), interleukin 1beta (IL-1beta) and tumor necrosis factor alpha (TNFalpha), which are known to stimulate the secretion of CINC/gro in the ovulatory process, and the effects of Keishi-bukuryo-gan with those of Toki-shakuyaku-san, which has been shown to have an effect on the ovary. We cultured whole ovarian dispersates from immature (3-week-old) female rats with Keishi-bukuryo-gan, Toki-shakuyaku-san and crude ingredients of Keishi-bukuryo-gan. The contents of CINC/gro, IL-1beta and TNFalpha in the cultured media were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: Keishi-bukuryo-gan stimulated the secretion of CINC/gro in a dose-dependent manner, and the secretion of CINC/gro into the culture medium increased significantly at concentrations of Keishi-bukuryo-gan of 10 and 100 microg/mL (P < 0.001). The stimulatory effect of Keishi-bukuryo-gan on the production of CINC/gro is significantly (P < 0.001) stronger than that of Toki-shakuyaku-san at the same concentrations of 100 microg/mL. In addition, Keishi-bukuryo-gan stimulated the secretion of IL-1beta in a dose-dependent manner, while it did not stimulate the secretion of TNFalpha even at a concentration of 100 microg/mL. Moutan Cortex, Paeoniae Radix and Persicae Semen, which are crude ingredients of Keishi-bukuryo-gan, enhanced the secretion of CINC/gro significantly (P < 0.01) in cultured whole ovarian dispersates. CONCLUSIONS: These results show that Keishi-bukuryo-gan can stimulate the secretion of CINC/gro as well as the production of IL-1beta and that this stimulatory effect of Keishi-bukuryo-gan was significantly stronger than that of Toki-shakuyaku-san in immature rat ovarian cell culture.     

The herbal medicine Unkei-to stimulates the secretion of a cytokine-induced neutrophil chemoattractant,CINC/gro,in the rat ovarian cell culture

PROBLEM AND METHOD OF STUDY: We investigated the ovulation-inducing effects of Unkei-to, a Japanese herbal medicine, in relation to the production of sex steroid hormones (17beta-estradiol and progesterone), cytokine-induced neutrophil chemoattractant (CINC/gro), interleukin-1beta (IL-1beta) and tumor necrosis factor alpha (TNF-alpha) in the rat ovarian cell culture. RESULTS: Unkei-to at a concentration of 100 microg/mL significantly stimulated the secretions of 17beta-estradiol and progesterone (P < 0.01) in cultured whole ovarian dispersates. Unkei-to also enhanced the secretion of CINC/gro in a dose-dependent manner, and the secretions of CINC/gro increased significantly at concentrations of 10 and 100 microg/mL (P < 0.01). These stimulatory effects of Unkei-to on steroidgenesis and CINC/gro production are very similar to those of another Japanese herbal medicine, Toki-Shakuyaku-san. In addition, Unkei-to significantly (P < 0.01) enhanced the secretions of both IL-1beta and TNF-alpha, which are known to stimulate the secretion of CINC/gro in the ovulatory process, at concentrations of 10 and 100 microg/mL. The stimulatory effect of Unkei-to at a concentration of 100 microg/mL on IL-1beta/was significantly (P < 0.01) lower than that of Toki-Shakuyaku-san, while the stimulatory effects of these two herbal medicines at a concentration of 100 microg/mL on TNF-alpha were similar. CONCLUSIONS: These results show that Unkei-to can stimulate ovarian steroidgenesis and the ovulatory process by inducing the secretion of CINC/gro with IL-1beta and TNF-alpha in vitro. Unkei-to has stimulatory effects on both steroidgenesis and the ovulatory process in the ovary as well as a stimulatory effect on the hypothalamus-pituitary axis, and it may be useful for treating patients with ovulatory disorders.     

Regulatory effects of interleukin-11 during acute lung inflammatory injury.

The role of interleukin-11 (IL-11) was evaluated in the IgG immune complex model of acute lung injury in rats. IL-11 mRNA and protein were both up-regulated during the course of this inflammatory response. Exogenously administered IL-11 substantially reduced, in a dose-dependent manner, the intrapulmonary accumulation of neutrophils and the lung vascular leak of albumin. These in vivo anti-inflammatory effects of IL-11 were associated with reduced NF-kappaB activation in lung, reduced levels of tumor necrosis factor alpha (TNF-alpha) in bronchoalveolar lavage (BAL) fluids, and diminished up-regulation of lung vascular ICAM-1. It is interesting that IL-11 did not affect BAL fluid content of the CXC chemokines, macrophage inflammatory protein-2 (MIP-2) and cytokine-inducible neutrophil chemoattractant (CINC); the presence of IL-11 did not affect these chemokines. However, BAL content of C5a was reduced by IL-11. These data indicate that IL-11 is a regulatory cytokine in the lung and that, like other members of this family, its anti-inflammatory properties appear to be linked to its suppression of NF-kappaB activation, diminished production of TNF-alpha, and reduced up-regulation of lung vascular ICAM-1.     

Rat Macrophage Inflammatory Protein-1α, a CC Chemokine, Acts as a Neutrophil Chemoattractant in Vitro and in Vivo

Recombinant rat macrophage inflammatory protein-1 (rMIP-1) at a concentration of 3 × 10^–8 M had strong neutrophil chemotactic activity, though the potency of rMIP-1 was less than that of cytokine-induced neutrophil chemoattractant (CINC)-1 at lower concentrations. In addition, rMIP-1 induced neutrophil chemotaxis in vivo when rMIP-1 was injected into the preformed air-pouch on the back of rats. The adhesion of rMIP-1-treated neutrophils to fibrinogen significantly increased, reaching a maximum adhesion at 10^–8 M. Stimulation of neutrophils with rMIP-1 induced a transient increase in intracellular free [Ca²⁺] dose-dependently. rMIP-1 still induced an increase in the intracellular [Ca²⁺] of rat neutrophils stimulated first with CINC-1, CINC-3 or C5a, suggesting that rat neutrophils have a specific receptor for rMIP-1. Supporting these findings, an additive increase in chemotactic potency was found when both rMIP-1 and CINC-1 were added to the lower wells of Boyden chamber in vitro. In addition, high levels of rMIP-1 were detected in the inflammatory site of air-pouch/carrageenan-induced inflammation in rats. Our results suggest that rMIP-1 acts as a neutrophil chemoattractant and, together with CINCs, plays an important role in infiltration of neutrophils into inflammatory sites in rats.     

Early Increase in Alveolar Macrophage Prostaglandin 15d-PGJ2 Precedes Neutrophil Recruitment into Lungs of Cytokine-Insufflated Rats

Early detection and prevention is an important goal in acute respiratory distress syndrome research. We determined the concentration of the anti-inflammatory 15-deoxy-Δ^12,14-prostaglandin-J2 (15d-PGJ2) and other components of the cyclopentenone prostaglandin cascade in relation to lung inflammation in cytokine (IL-1/LPS)-insufflated rats. We found that 15d-PGJ2 levels increase in the bronchoalveolar lavage (BAL) fluid of rats insufflated with cytokines 2 h before. BAL 15d-PGJ2 increases preceded neutrophil recruitment, lung injury, and oxidative stress in the lungs of cytokine-insufflated rats. 15d-PGJ2 was localized in alveolar macrophages that decreased following cytokine insufflation. 15d-PGJ2 may constitute an early biomarker of lung inflammation and may reflect an endogenous attempt to regulate ongoing inflammation in macrophages and elsewhere after cytokine insufflation.     

Induction of neutrophil infiltration by rat chemotactic cytokine (CINC) and its inhibition by dexamethasone in rats

In vivo effects of cytokine-induced neutrophil chemotactic factor (CINC) derived from rats on neutrophil infiltration were investigated using an air-pouch-type inflammation model in rats, and effects of dexamethasone on neutrophil infiltration induced by CINC was also examined in order to gain further insight into the mechanism of antiinflammutory activity of glucocorticoids. Injection of CINC into the air pouch made on the dorsum of rats induced a marked infiltration of neutrophils into the pouch fluid but not mononuclear cells and eosinophils during a 30-min interval after the injection. Maximum effect was induced at a dose of 1.4g/pouch. Treatment with dexamethasone 3 h before the injection of CINC suppressed the neutrophil infiltration in a dose-dependent manner, but no complete inhibition was observed. CINC injection into the air pouch of rats that had been sacrificed by bleeding in order to minimize neutroph il infiltration from blood stream also stimulated neutrophil infiltration into the pouch fluid when the carcass was incubated at 37C for 30 min, but the number of infiltrated neutrophils was about 35% of CINC-induced neutrophil infiltration in intact ruts. CINC-induced neutrophil infiltration in the carcass, which is supposed to be a reflection of neutrophil migration from extravascular space in subcutaneous tissues to pouch fluid, was not inhibited by dexamethasone treatment. Therefore, the inhibition of neutrophil infiltration by dexamethasone might be due to inhibition of the extravasation of peripheral neutrophils but not due to inhibition of neutrophil chemotaxis from subcutaneous extravascular space to pouch fluid. These findings suggest that clinical effects of steroidal antiinflammatory drugs on neutrophil infiltration in inflammatory disease is partly due to inhibition of neutrophil extravasation induced by preformed neutrophil chemotactic factors in the inflammatory site.     

Comparison of α2-Macroglobulin Synthesis by Juvenile vs. Mature Rats after Identical Inflammatory Stimulation

Synthesis of α₂-macoglobulin (α2M) by 3-week-old juvenile rats was compared to that of mature 7- and 11-week-old rats. Serum concentrations of α2M, interleukin (IL)-6- and cytokine-induced neutrophil chemoattractant (CINC)-1 were measured by enzyme-linked immunosorbent assay. The area under the concentration vs. time curve (AUC) for α2M was significantly different among the three groups. The synthesis of α2M increased in an age-dependent manner. No significant difference was observed for the AUC of IL-6, but that of CINC-1 in 3-week-old rats was significantly lower than that in 7- or 11-week-old rats. These results suggest that synthesis of α2M was increased in mature compared to juvenile rats, possibly due to differences in liver function. The maximum concentration of CINC-1 in 3-week-old rats was observed 6 h after turpentine oil injection. The serum concentrations of IL-6 and CINC-1 increased more quickly in juvenile rats than in mature rats after inflammatory stimulation.     

Evaluation of N-acetylcysteine treatment in acute pancreatitis-induced lung injury

Objective

Pulmonary complications are frequent during acute pancreatitis (AP). We investigate the effects of N-acetylcysteine (NAC) on lung injury in mild and severe AP.

Animals and treatment

Mild and severe AP was induced in rats by bile–pancreatic duct obstruction (BPDO) and infusion of 3.5?% sodium taurocholate (NaTc) into the bile-pancreatic duct, respectively. NAC (50?mg/kg) was given 1?h before and 1?h after AP.

Methods

Amylase activity was measured in plasma. Lungs were harvested for mRNA expression analysis of monocyte chemoattractant protein-1 (MCP-1), cytokine-induced neutrophil chemoattractant (CINC), P-selectin and intercellular adhesion molecule-1 (ICAM-1), myeloperoxidase (MPO) activity and histological examination.

Results

Hyperamylasemia was reduced by NAC in both AP models. NAC down-regulated MCP-1, CINC and P-selectin in BPDO- but not in NaTc-induced AP. Pulmonary insults did not vary in mild AP and were exacerbated in severe AP by NAC treatment. NAC reduced lung MPO activity in mild but not in severe AP.

Conclusions

Although NAC treatment down-regulated inflammatory mediators in lungs during AP it did not prevent leukocyte infiltration, which could be responsible for maintaining the lung injury. As a result, NAC aggravated the lung damage in severe AP and failed to exert beneficial effects in the mild disease model.     

Prophylactic effect of JTE-607 on LPS-induced acute lung injury in rats with CINC-1 inhibition

OBJECTIVE AND DESIGN: JTE-607, a multiple cytokine inhibitor, was evaluated in lipopolysaccharide (LPS)-induced acute lung injury in rats in vivo and in vitro. MATERIALS AND METHODS: LPS instillation into airways of rats was performed. JTE-607 at 3-30 mg/kg and dexamethasone at 3 mg/kg were administered intravenously at 10 min and 0 min for JTE-607, and 60 min for dexamethasone prior to the LPS instillation (n = 8). Cytokine-induced neutrophil chemoattractant (CINC)-1 level and myeloperoxidase (MPO) activity in lung were measured at 4 h after LPS instillation, and at 24 h for lung wet weight measurement and histological study. LPS-induced CINC-1 production by rat alveolar macrophages were also measured in vitro. RESULTS: JTE-607 and dexamethasone showed a significant reduction of increased CINC-1 level and MPO activity in lung after LPS treatment in vivo. Increased wet weight was also significantly inhibited. Histological studies revealed that JTE-607 and dexamethasone significantly inhibited LPS-induced accumulation of peribronchial neutrophils and eosinophils, and perivascular edema. JTE-607 and dexamethasone suppressed CfNC-1 synthesis by rat alveolar macrophages in vitro with IC50 values of 12.4 microM and 2.3 nM, respectively. CONCLUSIONS: These results indicate that JTE-607 has an inhibitory effect on LPS-induced rat lung inflammation in parallel with CINC-1 reduction. The effect of JTE-607 was suggested to be through direct inhibition of CINC-1 production from rat alveolar macrophages. JTE-607 may thus be efficacious in cytokine-mediated lung inflammation such as acute respiratory distress syndrome.     

Effects of Granulocyte Colony-Stimulating Factor (G-CSF) and Neutrophil Elastase Inhibitor (ONO-5046) on Acid-Induced Lung Injury in Rats

It has been suggested that neutrophils play an important role in acid-aspirated lung injury. We examined the effects of the high dose of granulocyte-colony stimulating factor (G-CSF), which is capable of increasing peripheral neutrophils, and a specific neutrophil elastase inhibitor (ONO-5046) on acid lung injury in rats. Animals were anesthetized and normal saline (NS, 2 mL kg⁻¹) or hydrochloric acid (HCl, 0.1 N 2 mL kg⁻¹) was then instilled into trachea. Thirty minutes before HCl instillation, G-CSF (150 μg kg⁻¹) was injected subcutaneously or ONO-5046 (10 mg kg⁻¹ h⁻¹) was infused continuously into the right jugular vein. Animals were ventilated during the experiments. Five hours after HCl or NS instillation, bronchoalveolar lavage fluid (BALF) and lung tissue samples were obtained. Total nuclear cell count, absorbance, albumin, tumor necrosis factor (TNF)-α, interleukin (IL)-6, cytokine-induced neutrophil chemoattractant (CINC), neutrophil elastase in BALF, wet-to-dry (W/D) ratio were measured. HCl aspiration markedly increased these values in BALF and W/D ratio. Both ONO-5046 and G-CSF attenuated the parameters increased by acid-induced lung injury in rats. The data suggests that neutrophils play an important role in acid-induced lung injury. However, high-dose G-CSF does not exacerbate acid-aspirated lung injury in rats, although this agent causes an increase in peripheral neutrophils.     

Functional immaturity of rat alveolar macrophages during postnatal development.

Alveolar macrophages (AM) are important in the regulation of immune responses in the lung, through their role as scavenger cells and through the production of many bioactive factors. Because in early infancy pulmonary infections are a recurrent problem, we studied the postnatal functional maturation of AM in a rat model. AM were isolated from rat lungs by bronchoalveolar lavage at several time intervals after birth and tested for their ability to ingest Escherichia coli in the presence of surfactant protein A (SP-A). Furthermore, their capacity to produce nitric oxide (NO) and interleukin-1 beta (IL-1 beta) after in vitro lipopolysaccharide (LPS) stimulation was analysed, as well as their capacity to downregulate proliferation of T cells from both mature and neonatal rats. SP-A-mediated phagocytosis of E. coli by AM was reduced in 14-day-old neonatal rats, as compared with mature rats (P < or = 0.05). Also the IL-1 beta production by rat AM after LPS stimulation was impaired at 14 days of age, as compared with IL-1 beta production by AM from mature rats (P < or = 0.05). In contrast, the LPS-induced NO production by rat AM as well as the capacity to inhibit T-cell proliferation were well developed at all ages tested. In conclusion, during postnatal development the rat AM is functionally immature, with respect to phagocytosis and secretion of inflammatory mediators. These differences may underly the enhanced susceptibility to pulmonary infections as found in human neonates.