The effect of extracts from ginger rhizome on inflammatory mediator production.

Compounds from rhizomes of Zingiber officinale, commonly called ginger, have been purported to have anti-inflammatory actions. We have used an in vitro test system to test the anti-inflammatory activity of compounds isolated from ginger rhizome. U937 cells were differentiated and exposed to lipopolysaccharide (LPS) from Escherichia coli (1 microg/ml) in the presence or absence of organic extracts or standard compounds found in ginger (6-, 8-, 10-gingerol or 6-shogaol) for 24 h. Supernatants were collected and analyzed for the production of prostaglandin E(2) (PGE(2)) and tumor necrosis factor alpha (TNF-alpha) by standard ELISA assays. Predominant compounds in the organic extracts were identified as 6-, 8- 10-gingerols and 6-, 8-, 10-shogaols. Organic extracts or standards containing gingerols were not cytotoxic, while extracts or standards containing predominantly shogaols were cytotoxic at concentrations above 20 microg/ml. Crude organic extracts of ginger were capable of inhibiting LPS induced PGE(2) (IC(50)<0.1 microg/ml) production. However, extracts were not nearly as effective at inhibiting TNF-alpha (IC(50)>30 microg/ml). Thirty three fractions and subfractions, prepared by column chromatography, were analyzed for bioactivity. Extracts containing either predominantly gingerols or shogaols (identified by HPLC) were both highly active at inhibiting LPS-induced PGE(2) production (IC(50)<0.1 microg/ml), while extracts that contained unknown compounds were less effective (IC(50)<3.2 microg/ml). Extracts or standards containing predominantly gingerols were capable of inhibiting LPS-induced COX-2 expression while shogaol containing extracts had no effect on COX-2 expression. These data demonstrate that compounds found in ginger are capable of inhibiting PGE(2) production and that the compounds may act at several sites.

The effect of turmeric extracts on inflammatory mediator production.

Major compounds of several commonly used botanicals, including turmeric, have been purported to have anti-inflammatory actions. In order to test the anti-inflammatory activity of compounds isolated from rhizomes of Curcuma longa L. (Zingiberaceae), we have established an in vitro test system. HL-60 cells were differentiated and exposed to lipopolysaccharide (LPS) from Escherichia coli (1 microg/ml) in the presence or absence of botanical compounds for 24 h. Supernatants were collected and analyzed for the production of tumor necrosis factor alpha (TNF-alpha) and prostaglandin E2 (PGE2) using standard ELISA assays. Water-soluble extracts were not cytotoxic and did not exhibit biological activity. Organic extracts of turmeric were cytotoxic only at concentrations above 50 microg/ml. Crude organic extracts of turmeric were capable of inhibiting LPS-induced TNF-alpha (IC50 value = 15.2 microg/ml) and PGE2 (IC50 value = 0.92 microg/ml) production. Purified curcumin was more active than either demethoxy- or bisdemethoxycurcumin. Fractions and subfractions of turmeric extracts collected via preparative HPLC had differing biological activity, ranging from no activity to IC50 values of < 1 microg/ml. For some fractions, subfractionation resulted in a loss of activity, indicating interaction of the compounds within the fraction to produce an anti-inflammatory effect. A combination of several of the fractions that contain the turmeric oils was more effective than the curcuminoids at inhibiting PGE2. While curcumin inhibited COX-2 expression, turmeric oils had no effect on levels of COX-2 mRNA.

Sodium arsenite enhances AP-1 and NFkappaB DNA binding and induces stress protein expression in precision-cut rat lung slices.

Arsenic is a known human carcinogen. These studies were designed to examine the impact of low arsenite concentrations on immediate early gene expression in precision-cut rat lung slices. Precision-cut lung slices are a versatile in-vitro system for toxicity studies, as they preserve the architecture and cellular heterogeneity of the lung. Since 0.1-100 microM arsenite did not compromise slice viability at 4 hours, effects of arsenite on the expression of c-jun/AP-1, NFkappaB, HSP 32, HSP 72, HSP 60, and HSP 90 were studied, using these concentrations of arsenite at 4 h. Nuclear c-jun was increased by 10 and 100 microM arsenite, while NFkappaB was not affected. Gel-shift assays indicated that 10 microM arsenite resulted in an enhanced DNA-binding activity of both AP-1 and NFkappaB. Confocal microscopic analysis of AP-1 indicated nuclear localization of this transcription factor, mainly in type-II epithelial cells and alveolar macrophages. Nuclear localization of NFkappaB was lower than that observed for AP-1, while most of the NFkappaB was localized to cytoplasm of type-II epithelial cells and alveolar macrophages. HSP 32 was increased by 1.0 and 10 microM arsenite, while HSP 72 was increased by only 100 microM arsenite. HSP 60 and HSP 90 were not changed by arsenite. These studies indicate that noncytotoxic concentrations of arsenite are capable of affecting signal transduction pathways and gene expression in the lung.

Adverse respiratory effects following overhaul in firefighters.

Overhaul is the stage in which firefighters search for and extinguish possible sources of reignition. It is common practice not to wear respiratory protection during overhaul. Fifty-one firefighters in two groups, 25 without respiratory protection and 26 wearing cartridge respirators, were monitored for exposure to products of combustion and changes in spirometric measurements and lung permeability following overhaul of a structural fire. Testing at baseline and 1 hour after overhaul included forced vital capacity (FVC), forced expiratory volume in one second (FEV1), serum Clara cell protein (CC16), and serum surfactant-associated protein A (SP-A). Overhaul increased CC16 in both groups, indicating increased alveolarcapillary membrane permeability. Contrary to expectations, SP-A increased and FVC and FEV1 decreased in the firefighters wearing cartridge respirators. Changes in FEV1, CC16, and SP-A were associated with concentrations of specific products of combustion or carboxyhemoglobin levels. Firefighter exposures during overhaul have the potential to cause changes in spirometric measurements and lung permeability, and self-contained breathing apparatus should be worn during overhaul to prevent lung injury.

Rapid reduction of intracellular glutathione in human bronchial epithelial cells exposed to occupational levels of toluene diisocyanate.

Toluene diisocyanate (TDI) is a recognized chemical asthmogen, yet the mechanism of this toxicity and the molecular reactions involved have not been elucidated. We have previously shown that TDI vapor forms adducts with the apical surface of the respiratory epithelium, and that it colocalizes with ciliary tubulin. In vitro, we have shown rapid reaction of TDI with glutathione (GSH) and transfer of the bisGS-TDI adduct to a sulfhydryl-containing major histocompatibility complex peptide. This study sought to determine if intracellular GSH is altered following exposure to TDI. We used the dye CellTracker Green (chloromethylfluorescein, CMFDA) for detection of glutathione. One-day and 6-day air-liquid cultures of human bronchoepithelial cells (HBE) were exposed to 20-100 ppb TDI vapor for 5, 15, or 30 min. Cells were subsequently imaged using a confocal microscope. Both 1- and 6-day cultures showed a decrease in intensity of the thiol staining as a function of the TDI exposure dose. Doses as low as 20 ppb, the current permissible exposure limit (PEL) to TDI, resulted in rapid (within 5 min) decreases in fluorescence. The decreased fluorescence was not due to cytotoxicity or decrease in either esterase or glutathione-S-transferase activity, enzymes necessary for activation of the fluorescence of CMFDA. The decrease in glutathione levels was verified using another fluorescent label, ThioGlo(TM) 1, and cell extracts. In addition, the mucus produced by 6-day air-liquid interface HBE cells in response to TDI exposure appeared to be protective, as HBE cells underlying mucus retained more fluorescence than did cells in the same cultures that were not covered with mucus. These results, along with previous data, strongly suggest that TDI enters pulmonary cells and reacts rapidly with intracellular GSH, and that this can occur at the current PEL of 20 ppb. This rapid reaction suggests the importance of cellular thiols in TDI-induced pulmonary disease.

Short-term pulmonary response to inhaled JP-8 jet fuel aerosol in mice.

B6.A.D. (Ahr(d)/Nat(s)) mice were utilized to investigate the short-term pulmonary response to JP-8 jet fuel (JP-8) aerosol inhalation. Mice were nose-only exposed to atmospheres of 0 to 118 mg/m3 for 1 h/d over a period of 7 days to further test the hypothesis that JP-8 concentrations below the permissible exposure level (PEL) of 350 mg/m3 will induce lung injury. At 24 to 30 hours after the final exposure, pulmonary function and respiratory permeability were measured on anesthetized mice and then randomly assigned for bronchoalveolar lavage or histopathology. Bronchoalveolar lavage fluid (BALF) was analyzed for total protein, lactic dehydrogenase (LDH), N-acetyl-beta-D-glucosaminidase (NAG), and cytology. Respiratory permeability increases were observed following doses of 48 and 118 mg/m3 and were supported by concomitant BALF increases in total protein and LDH. Conversely, NAG and alveolar macrophage levels decreased following the same exposure concentrations. Morphological lung injury was characterized by the targeting of bronchiolar epithelium and consisted of perivascular edema, Clara cell vacuolization, and necrosis. Alveolar injury included sporadic pulmonary edema, intra-alveolar hemorrhage, and alterations in type II epithelial cells. These results indicate that repeated inhalation of aerosolized JP-8 induces physiological, biochemical, cellular, and morphological lung injury. This study also provides evidence for the reevaluation of the 350 mg/m3 PEL for more volatile petroleum distillates with regard to respirable aerosols.

Nedocromil sodium inhibits canine adenovirus bronchiolitis in beagle puppies.

Nedocromil sodium is a nonsteroidal anti-inflammatory drug used to control asthmatic attacks. Our hypothesis is that nedocromil sodium inhibits virus-induced airway inflammation, a common trigger of asthma. We nebulized nedocromil sodium into beagle dogs (n = 10, mean +/- SEM ages: 149 +/- 13 days) before and after inoculation with canine adenovirus type 2 (CAV2). Control dogs (n = 10) received saline aerosols and were either infected with CAV2 (Sal/CAV2, n = 7, mean +/- SEM ages: 140 +/- 11 days) or were not infected (Sal/Sal, n = 3, ages: 143 +/- 0 days). All dogs were anesthetized with choralose (80 mg/kg i.v.), intubated, and mechanically ventilated. Pulmonary function tests and bronchoalveolar lavage (BAL) were performed using standard techniques. Pulmonary function tests revealed no significant change between the nedocromil sodium and non-nedocromil-treated groups. The percentage of infected bronchioles was quantitated as the number of inflamed airways of 40 bronchioles examined times 100 for each dog. Nedocromil-treated dogs had significantly (p < 0.05) less mucosal inflammation (mean +/- SEM, 39% +/- 5%), epithelial denudation (36% +/- 5%), and BAL neutrophilia (11 +/- 3) than did Sal/CAV2 dogs (51% +/- 6%, 57% +/- 4%, and 33% +/- 8%, respectively). We concluded that pretreatment with nedocromil sodium aerosols attenuated CAV2-induced airway inflammation in these beagle puppies.

Functional alterations of alveolar macrophages subjected to smoke exposure and antioxidant lazaroids.

Acute inhalation of diesel fuel-polycarbonate plastic (DFPP) smoke causes severe lung injury, leading to acute respiratory distress syndrome (ARDS) and death. It has been reported that the initiation of acute lung injury is associated with the activation of pulmonary alveolar macrophages (PAM). To further explore the pathogenesis, alveolar macrophages (AM) of New Zealand rabbits ventilated and exposed to a 60 tidal volume of DFPP smoke in vivo were recovered at 1 h post-smoke. Smoke exposure induced significant increases in both mRNA and protein levels for PAM tumor necrosis factor-alpha (TNF-alpha), when compared to smoke control. Smoke also induced a biphasic response (inhibited at 2 h, enhanced at 24 h after cell isolation) in the production of superoxide (O2-) by PAM. However, aerosolized lazaroid, U75412E (1.6 mg/kg body weight), significantly attenuated smoke-induced expression in AM TNF-alpha at the protein level but not at the mRNA level, and smoke-induced changes in AM production of O2-. This study suggests that highly expressing AM TNF-alpha following smoke may be a key contributor to the cascade that establishes an acute injury process and exacerbates oxidant-derived cell injury. Whereas, the lazaroid may ameliorate smoke-induced lung injury by attenuating AM TNF-alpha release, in addition to its primary antioxidative mechanism.

Toluene diisocyanate colocalizes with tubulin on cilia of differentiated human airway epithelial cells.

Toluene diisocyanate (TDI), a highly reactive industrial chemical with widespread use in the manufacture of polyurethane and plastics, is the leading cause of occupational asthma associated with chemical exposure. We report the effects of TDI vapor (20, 100, 500, 1000 ppb) in vitro on differentiated human bronchial epithelial cells. Increased mucus was observed by electron microscopy at all TDI concentrations. Cytotoxicity, as evidenced by cell pyknosis and DNA fragmentation, was detected following a 30-min exposure to TDI concentrations of 100 ppb or higher. At 1000 ppb, transepithelial resistance was lost. Using confocal microscopy and double staining, TDI was found colocalized with ciliary tubulin in cultures that had been exposed to 20 and 100 ppb. These findings are the first to identify TDI binding to human pulmonary epithelial cells and indicate extensive binding to the cilia of differentiated epithelial cells. The in vivo implications of these findings include decreased ciliary movement and longer retention of TDI and hence increased exposure. Altered cytoskeletal-derived signal transduction may be a consequence of tubulin involvement. The effects of such changes on respiratory sensitization remain to be explored.

Early alterations of lung injury following acute smoke exposure and 21-aminosteroid treatment.

In a simulated fire-related smoke exposure protocol, New Zealand white rabbits were utilized to investigate the potential effects of the 21-aminosteroid (lazaroid) analog U75412E on the early events of acute lung injury. Inhalation of a total of 1.6 mg/kg U75412E aerosolized at a rate of 1.53 mg/min at 0.5 hr after smoke exposure significantly attenuated the extent of lung injury at 1 hr, as evidenced by decreased bronchoalveolar lavage (BAL) concentration of total protein, 6-keto-prostaglandin F1-alpha, and blood gas defect. Histopathologic examination demonstrated that the lazaroid significantly attenuated smoke-induced lung injury as evidenced by a decrease in wet lung/body weight ratio, necrosis, and sloughing of airway epithelial cells. Electron microscopy showed that the lazaroid decreased smoke-induced interstitial edema and the vacuolization of alveolar type II epithelium (21.6 +/- 9.7 vs 8.5 +/- 3.6 vacuoled blebs/cell, smoke only vs smoke + lazaroid). However, U75412E did not attenuate smoke-induced changes in BAL concentration of tumor necrosis factor-alpha, total cell count, and granulocyte percentage. These observations suggest that U75412E may exert its action through cooperative mechanisms, such as the modulation of arachidonic acid metabolism, in addition to its characterized antioxidative effects.

1,2-Dichlorobenzene-induced lipid peroxidation in male Fischer 344 rats is Kupffer cell dependent.

1,2-Dichlorobenzene (1,2-DCB) is a potent hepatotoxicant in male Fischer 344 (F344) rats and previous studies have suggested that reactive oxygen species may play a role in the development of hepatotoxicity. Since reactive oxygen species can damage lipid membranes, this study was conducted to determine the extent of lipid peroxidation after administration of 1,2-DCB by immuno-histochemical analysis of 4-hydroxynonenal (4-HNE) protein adduct formation in liver and conjugated diene formation in liver and serum. The contribution of Kupffer cells to the lipid peroxidation was also investigated. Male F344 rats were administered 1,2-DCB (3.6 mmol/kg i.p. in corn oil) and killed at selected times between 3 and 48 h. Time course studies revealed the greatest abundance of 4-HNE protein adducts in the centrilobular regions of the liver 24 h after 1,2-DCB administration, with much lower levels at 16 h. Adducts were present in necrotic and vacuolized centrilobular hepatocytes of 1,2-DCB treated rats but not in livers of controls. Further, conjugated dienes were significantly increased in liver and serum 16 and 24 h after 1,2-DCB administration, peaking at 24 h. These data correlated with hepatocellular injury, determined by serum alanine aminotransferase activity and histopathological evaluation, which was markedly elevated within 16 h and peaked at 24 h. When rats were pretreated with gadolinium chloride (GdCl3; 10 mg/kg i.v. 24 h prior to 1,2-DCB), an inhibitor of Kupffer cells, hepatotoxicity was decreased by 89 and 86%, at 16 and 24 h, respectively. Conjugated diene concentrations were decreased to control values at these times after 1,2-DCB administration. Moreover, no 4-HNE protein adducts were detected in livers of 1,2-DCB-treated rats pretreated with GdCl3. Finally, Kupffer cells isolated from 1,2-DCB-treated rats produced significantly more superoxide anion than Kupffer cells isolated from vehicle controls. These data, along with previous findings, suggest that lipid peroxidation associated with 1,2-DCB is mediated in part by Kupffer cell-derived reactive oxygen species.

Modulation of Kupffer cell and peripheral blood monocyte activity by in vivo treatment of rats with all-trans-retinol.

Previous studies have shown that large doses of vitamin A potentiate chemical-induced liver injury and that the Kupffer cell is directly involved in this potentiation. Therefore, these studies were designed to determine if Kupffer cells isolated from vitamin A treated male Sprague-Dawley rats (75 mg/kg/day for 3-7 days as all- trans-retinol) had altered activity and function. Respiratory activity of Kupffer cells isolated from rats treated with vitamin A for 3 to 7 days markedly increased. Similarly, phagocytic activity was significantly elevated (up to 9-fold) after exposure to vitamin A for 3 to 7 days. Production of reactive oxygen species, measured by luminol-enhanced chemiluminescence of Kupffer cells isolated after 7 days of vitamin A exposure, was significantly higher than that of control cells when stimulated with opsonized zymosan. Also, the release of superoxide anion by individual Kupffer cells isolated from vitamin A treated rats was nearly three times greater than that of control cells. Basal production of tumor necrosis factor-alpha (TNF-alpha) and prostaglandin E2 (PGE2) production were significantly elevated in Kupffer cells isolated from rats treated with vitamin A. Lastly, peripheral blood monocytes (PBMC) isolated from rats treated with vitamin A for 7 days had a significantly greater respiratory activity, as well as TNF-alpha and PGE2 production, than PBMC isolated from control rats. Our data suggest that large doses of vitamin A enhance both Kupffer cell and PBMC function. Upregulation of the activity by these phagocytic cells may play a role in the vitamin A potentiation of chemical-induced liver injury.

Fractal analysis of lung alveoli during the acute phase vs. repair phase of an adenoviral infection in canines.

Acute viral respiratory infections are commonly associated with alterations in lung growth. Recently, fractal techniques have been demonstrated to show changes in alveolar perimeter after canine adenovirus type 2 (CAV2) infection in a beagle puppy model. In the present study, we investigated whether the fractal dimension (Df) of the alveolar perimeter was changed in the acute phase (2-3 weeks after inoculation, 131d CAV2 group) or during the recovery phase (approximately 22 weeks after inoculation, 235d CAV2 group) after a single bout of CAV2. There were sham CAV2 groups, 130d and 238d controls, corresponding to the CAV2 groups. The Df of alveolar perimeter length was significantly increased in the 235d CAV2 puppies compared to all of the other beagle puppy groups. On the other hand, the fractal dimensions for alveolar perimeter length for the other beagle puppy groups were very similar. In a related human study of patients (age range of 25 h to 19 y, N = 11), who died of non-respiratory causes, showed no consistent change in Df of alveolar perimeter length with normal lung growth and development. We conclude that fractal analysis of alveolar perimeter length can be used as an index of permanent lung injury after insult to the growing lungs.

A free radical scavenger (Lazaroid U75412E) attenuates tumor necrosis factor-alpha generation in a rabbit model of smoke-induced lung injury.

The lazaroid (21-aminosteroid) analogue U75412E was evaluated in rabbits exposed to diesel fuel-polycarbonate plastic smoke. Inhalation of total of 4.6 mg U75412E aerosolized at a rate of 1.53 mg/min for 3 min before or after smoke significantly prevented or limited the extent of alveolar hypoventilation, interstitial edema, and tumor necrosis factor-alpha (TNF-alpha) by pulmonary alveolar macrophages (PAM) ex vivo observed at 2 h. The smoke-induced changes in wet lung/body weight ratios and the production of superoxide (O2-) by PAM ex vivo were also attenuated by the drug treatment after smoke exposure (p < 0.05). This study suggests that lazaroids may ameliorate the oxygen-radical-initiated cytokine processes and inflammation cascade as a result of the smoke insult.

Human alveolar fractal dimension in normal and chronic obstructive pulmonary disease subjects.

We have determined that fractal analysis of the alveolar perimeter (Df) changes with aging in human lung tissue in twenty-nine patients, age range of 25 hours to 76 years, who died of non-respiratory related causes. There was a very significant difference (p = 0.0004) in Df between the young (less than 16 years old, N = 9, mean Df of 1.047 [0.01]) and adult (greater than 16 years old, N = 20, mean Df of 1.093 [0.013]) groups. Furthermore, there was a significant difference in Df between the Adult group and the group of patients who died of chronic obstructive pulmonary disease (COPD, N = 10) (p = 0.012). Additionally, the Df values for the COPD and cystic fibrosis (CF, N = 5) groups were virtually identical; 1.061 and 1.070, respectively. Regression analysis showed a significant (p = 0.0041) exponential relationship with a correlation coefficient of 0.59 between aging and Df. We have demonstrated that the correlation between Df and aging in humans is an exponential function and that the end-stage pulmonary diseases of COPD and CF decrease Df.

The prophylactic effects of U75412E pretreatment in a smoke-induced lung injury rabbit model.

The effects of the lazaroid analogue U75412E (21-[4-(3-ethylamino-2-pyridinyl)-1-piperazinyl]-16 alpha-methylpregna-1,4,9]-(11)-triene-3,20-dione) were examined in an acute lung injury rabbit model. Standard doses of 0, 8 and 16 mM U75412E were aerosolized and ventilated into the lungs for 3 min. via an endotracheal tube. A 60 tidal volume dose of diesel fuel-polycarbonate plastic smoke was then instilled, followed by mechanical ventilation for one hour. Pretreatment with 16 mM U75412E significantly increased blood PaO2 and pH values, and decreased blood PaCO2 as compared to smoke only exposures. It also significantly decreased the total cell counts and granulocytes in bronchoalveolar lavage fluid, and the ability of pulmonary alveolar macrophages to produce tumour necrosis factor-alpha in vitro after cell isolation and culture. Histopathology indicated that 16 mM U75412E pretreatment attenuated increases in wet lung/body weight ratios, inflammatory focus, and interstitial oedema associated with smoke insult. In summary, U75412E pretreatment may possess the potential to improve acute smoke-induced lung injury, in part, through modulation of tumour necrosis factor-alpha production from pulmonary alveolar macrophages.

Neutral endopeptidase (NEP) and its role in pathological pulmonary change with inhalation exposure to JP-8 jet fuel.

Through a simulated flightline exposure protocol, Fischer 344 rats (F344) were subjected to an aerosol/vapor mix of the military jet fuel, JP-8. Previous studies with this model of lung injury have revealed significant increases in pulmonary resistance, increased alveolar clearance of 99mTcDTPA, and a decrease in bronchoalveolar lavage fluid (BALF) concentration of the neuropeptide substance P (SP). Exposures to JP-8 were nose-only and for one hour daily. Six groups of Fischer 344 rats were exposed for 7, 28, or 56 days at two JP-8 concentrations (low dose = 469-520 mg/m3/hr, high dose = 814-1263 mg/m3/hr). Exposed groups were matched with longitudinal controls. In response to JP-8 inhalation, exposure animals demonstrated a dose-dependent as well as duration-determined reduction in BALF SP concentration. Both JP-8 concentrations caused significant pathological changes in lower pulmonary structures.

Inhalation exposure to JP-8 jet fuel alters pulmonary function and substance P levels in Fischer 344 rats.

In a simulated military flightline exposure protocol, Fischer 344 rats (F344) were used to investigate the pulmonary effects of JP-8 jet fuel inhalation. Exposures were nose only and for 1 h daily. Groups were exposed for 7 days (7D) or 28 days (28D). Each exposure group had a matched longitudinal control group (LC7 and LC28). Exposure concentrations of 520 mg m-3 caused an increase in dynamic compliance after 7 days of exposure, but compliance changes were not seen with continued exposure (28D, 495 mg m-3). Pulmonary resistance was increased in both 7- and 28-day JP-8-exposed groups. Changes in pulmonary function were accompanied by a decrease in substance P concentrations from the bronchoalveolar lavage fluid (BALF). No significant change was observed in BALF levels of 6-keto-PGF1 alpha, the stable metabolite of prostacyclin, which is a marker of endothelial cell function. The JP-8-exposed rats gained significantly less weight during the study period than the LC7 and LC28 groups, and the lungs of the 7D group were heavier by wet lung/body weight ratio (WtL/WtB). Alveolar clearance of technetium-labelled diethylenetriamine pentaacetate ([99mTc]DTPA) was increased in jet fuel-exposed groups. Light microscopy showed no pathological evidence of lung injury. Recovery from the early pulmonary effects of JP-8 inhalation occurred with continued exposure, as seen by recovery of pulmonary compliance and WtL/WtB.

Locomotive forces produced by single leukocytes in vivo and in vitro.

We report here the first time-resolved measurements of the forces produced during the migration of single leukocytes in vivo and in vitro. Pulmonary macrophages from hamsters and mice, in vitro, and Nembutal (pentobarbital sodium)-anesthetized hamster neutrophils, in vivo, generated maximum locomotive forces ranging from 1.9 to 10.7 nN or tenths of microdynes. Force production was periodic and correlated with the length of the leading lamellipod but not with generalized cell ruffling. Although the extension of the leading lamella is critical to locomotive force generation, these direct measurements suggest that lamellar extension may not arise from the same contractile processes driving forward motion of the cell mass. Indeed, cell ruffling, lamellar extension, and locomotive force generation may be differentially controlled and have different origins. This technique may be extended to test numerous hypotheses of how these and other nonmuscle cells crawl.

Changes in lung permeability correlate with lung histology in a chronic exposure model.

In a simulated military flight-line exposure protocol, the effects of JP-8 jet fuel exposure on lung epithelial permeability were evaluated in male Fischer 344 rats (F344). Exposures were nose-only and for one hour daily. Groups were exposed for 7, 28, and 56 days. A protocol for administering a low dose (500mg/m3/hr) and a high dose (813-1094mg/m3/hr) of JP-8 jet fuel was used. Longitudinal sham-exposure groups (no jet fuel) for 7, 28, and 56 days were included in the protocol. Lung epithelial permeability was measured by clearance of technetium-labeled diethylenetriamine pentaacetate (99mTcDTPA, molecular weight = 492 daltons, physical half-life = 6.02 hours). The percent clearance of 99mTcDTPA per minute was calculated. Alveolar epithelial clearance for JP-8-exposed rats was dependent on both exposure concentration and duration. It was noted that at low-dose exposure concentrations alveolar epithelial clearance of 99mTcDTPA returned to low levels (LD56 = 1.09% per min; LC56 = 0.98% per min), suggesting recovery as evidenced by microscopic exam. The corresponding 56-day high-dose group (n = 10) had a significantly higher (p < 0.05) value of 2.25% per minute. The 28-day low-dose (n = 15) and high-dose (n = 20) groups had clearance values that were significantly increased from their longitudinal control group (n = 17). The alveolar epithelial permeability values were 2.51, 1.95, and 1.20, respectively. The seven-day longitudinal control, low-dose, and high-dose groups had alveolar permeability values of 1.57, 2.16, and 2.07, respectively. The lung histology correlated with the clearance values. Electron micrographs showed that all groups had interstitial edema resulting from endothelial damage. There was apparent thickening of the alveolar septa, and alveolar macrophages were activated in all groups. Lung permeability data, as determined by 99mTcDTPA alveolar clearance, indicated that lung injuries peaked at 28 days of jet fuel exposure, and this finding corresponded with the histology data. There was a discrepancy in the seven-day group between the number of cells and the 99mTcDTPA clearance values. The HD7 group had a total cell count significantly higher than all other groups, but the 99mTcDTPA clearance values in that group were not significantly different from that of any other group.