Enhanced allergic airway disease in old mice is associated with a Th17 response.

BACKGROUND: The prevalence of asthma in the elderly is increasing and associated with higher mortality than in children or young adults. However, the effects of age on the development and character of allergic asthma have been understudied. It has been suggested that mixed Th2/Th17 responses cause more severe forms of asthma, but the role of Th17 response in allergic airway disease and aging is not well understood. OBJECTIVE: To investigate age-dependent characteristics and Th17 immune response in allergic airway disease in a murine house dust mite (HDM)-allergen model. METHODS: Twelve-week-old and 15-month-old male BALB/c mice were sensitized and challenged with HDM. Bronchoalveolar lavage fluid (BALF), airway inflammation and hyperresponsiveness (AHR), serum immunoglobulin and splenic T cells were assessed. Age-related T cell activation was analyzed in a co-culture with bone marrow-derived dendritic cells (BMDC) and splenic CD4(+) T cells from young and old mice. RESULTS: Features of allergic airway disease such as mucous cell hyperplasia, infiltration of airway eosinophils and lymphocytes, Th2 cytokine expression and serum IgG1 levels were greater in old compared to young mice. In contrast to the more marked inflammatory/immune responses to HDM in old mice, AHR was greater in young HDM-treated mice. Only the old mice developed airway neutrophil infiltration and a Th17 immune response upon HDM exposure, with increases in BALF cytokines IL-17A and KC, and Th17 cytokine producing T cells in the spleen. Stimulation of CD4(+) T cells and BMDC co-cultures with HDM, resulted in an enhanced Th17 cytokine response in cells isolated from old mice. CONCLUSIONS AND CLINICAL RELEVANCE: Our findings in mice suggest that the severity and character of allergic airway disease are age dependent, with a bias towards a Th17 immune response with aging. Elderly, asthmatics may be prone to develop severe allergic airway inflammation with a mixed Th2/Th17 immune response.

Gamma-tocopherol prevents airway eosinophilia and mucous cell hyperplasia in experimentally induced allergic rhinitis and asthma.

BACKGROUND: Traditional therapies for asthma and allergic rhinitis (AR) such as corticosteroids and antihistamines are not without limitations and side effects. The use of complementary and alternative approaches to treat allergic airways disease, including the use of herbal and dietary supplements, is increasing but their efficacy and safety are relatively understudied. Previously, we have demonstrated that gamma-tocopherol (gammaT), the primary form of dietary vitamin E, is more effective than alpha-tocopherol, the primary form found in supplements and tissue, in reducing systemic inflammation induced by non-immunogenic stimuli. OBJECTIVE: We used allergic Brown Norway rats to test the hypothesis that a dietary supplement with gammaT would protect from adverse nasal and pulmonary responses to airway allergen provocation. METHODS: Ovalbumin (OVA)-sensitized Brown Norway rats were treated orally with gammaT before intranasal provocation with OVA. Twenty-four hours after two challenges, histopathological changes in the nose, sinus and pulmonary airways were compared with gene expression and cytokine production in bronchoalveolar lavage fluid and plasma. RESULTS: We found that acute dosing for 4 days with gammaT was sufficient to provide broad protection from inflammatory cell recruitment and epithelial cell alterations induced by allergen challenge. Eosinophil infiltration into airspaces and tissues of the lung, nose, sinus and nasolacrimal duct was blocked in allergic rats treated with gammaT. Pulmonary production of soluble mediators PGE(2), LTB(4) and cysteinyl leukotrienes, and nasal expression of IL-4, -5, -13 and IFN-gamma were also inhibited by gammaT. Mucous cell metaplasia, the increase in the number of goblet cells and amounts of intraepithelial mucus storage, was induced by allergen in both pulmonary and nasal airways and decreased by treatment with gammaT. CONCLUSIONS: Acute treatment with gammaT inhibits important inflammatory pathways that underlie the pathogenesis of both AR and asthma. Supplementation with gammaT may be a novel complementary therapy for allergic airways disease.

Toxicant-induced oxidative stress in cancer.

The article highlighted in this issue is "The Role of Oxidative Stress in Indium Phosphide-Induced Lung Carcinogenesis in Rats" by Barbara C. Gottschling, Robert R. Maronpot, James R. Hailey, Shyamal Peddada, Cindy R. Moomaw, James E. Klaunig, and Abraham Nyska (pp. 28-40). The article integrates a traditional pathologic study of toxicant-induced pulmonary carcinogenesis with an immunohistologic assessment of oxidative stress, thereby determining a potential mechanism of action of a toxicant, specifically indium phosphide.

Effects of ozone and endotoxin coexposure on rat airway epithelium: potentiation of toxicant-induced alterations.

Tropospheric ozone is the major oxidizing component in photochemical smog and is one of the most pervasive problems to human health of the criteria air pollutants for which the National Ambient Air Quality Standards have been designated by the Clean Air Act. Although many adverse health effects of ozone exposure have been documented in both humans and laboratory animals, controversy surrounds the establishment and implementation of ozone standards set forth by the U.S. Environmental Protection Agency. Because people are commonly exposed to more than one air pollutant at a time, studies that examine coexposures to airborne materials may be more relevant for assessing their risks to human health. Airborne biogenic substances such as pollens, spores, and bacterial products are ubiquitous in the environment, and when inhaled can cause adverse respiratory symptoms. One such biogenic agent, bacterial endotoxin, is a potent stimulus of airway inflammation and is a ubiquitous airborne contaminant commonly found in domestic, agricultural, and industrial settings. Little is known about the interaction of exposures to biogenic substances and criteria air pollutants such as ozone. In the last few years we have performed a series of studies in rodents that examined the biologic responses of the respiratory epithelium after airway exposures to both endotoxin and ozone. When exposed to ozone (0.5 ppm 8 hr/day for 3 days), Fischer rats develop lesions in the nasal transitional epithelium, whereas intranasal instillation of endotoxin (20 microg) elicits epithelial lesions in the respiratory epithelium of the nose and conducting airways. Our studies were designed to examine how exposure to one toxicant may affect the airway epithelial lesions induced by the other toxicant. We investigated the potential role of acute inflammation in the enhancement of airway epithelial lesions after exposure of these two toxicants in neutrophil-sufficient and neutrophil-deficient rodents. A summary of these results indicates that epithelial and inflammatory responses to coexposure of these two pollutants are greater than those elicited by either agent alone. Interestingly, each toxicant enhances the epithelial alterations induced by the other. Furthermore, the synergistic effects elicited by coexposure to ozone and endotoxin are mediated partly by neutrophils. These studies provided some new insights into how inhaled co-pollutants interact to initiate and promote alterations of airway epithelium. Further studies with these and other air pollutants will help define their true risk to human health.

Endotoxin enhancement of ozone-induced mucous cell metaplasia is neutrophil-dependent in rat nasal epithelium.

Ozone, the primary oxidant gas in photochemical smog, causes neutrophilic inflammation and mucous cell metaplasia (MCM) in the nasal transitional epithelium (NTE) of rats and monkeys. Bacterial endotoxin is another common airborne agent that induces acute neutrophilic inflammation, but not MCM, in NTE. It does, however, enhance ozone-induced MCM in rat nasal airways (Fanucchi et al., 1998, Toxicol. Appl. Pharmacol. 152, 1-9). In the present study, F344 rats exposed to filtered air or 0.5 ppm ozone (8 h/day for 3 days) were intranasally instilled with sterile saline or 100 microg endotoxin 24 h and 48 h after the third ozone exposure. To determine the role of neutrophilic inflammation in endotoxin-induced potentiation of the MCM caused by ozone, half of the rats were depleted of circulating neutrophils prior to saline or endotoxin instillations. Rats were killed 6 h or 3 days after the last intranasal instillation, and nasal tissues were processed for (1) light microscopy and morphometric analysis to determine the number of infiltrating neutrophils and the volume amount (density) of stored mucosubstances in the NTE, and (2) quantitative RT-PCR analysis of steady-state mucin gene (rMuc-5AC) mRNA levels in the NTE. Endotoxin induced a transient influx of neutrophils in both air- and ozone-exposed rats that was completely blocked by neutrophil depletion. Endotoxin increased rMuc-5AC mRNA levels in the NTE of ozone-exposed rats. Neutrophil depletion, however, had no effect on endotoxin-induced upregulation of mucin gene mRNA levels. Endotoxin enhanced the ozone-induced increase in stored mucosubstances (4-fold increase), but only in neutrophil-sufficient rats. These data indicate that endotoxin enhancement of ozone-induced upregulation of rMuc-5AC mRNA levels is neutrophil-independent, while its effects on intraepithelial production and storage of mucus glycoproteins is dependent on the presence of neutrophils.

Comparison of methods for measuring root and mucogingival sensitivity.

OBJECTIVE: The aim of this study was to compare the variability of measurements of root and mucogingival sensitivity over a 24-hour period. STUDY DESIGN: Sixteen individuals (46.8 +/- 3.2 years old) were randomly tested for pain thresholds with calibrated electrical stimulation of the root and adjacent mucosa (electric pulp tester), pressure on mucosa (pressure-sensitive probe), and cold on the root (experimental thermocoupler probe) at baseline and after 4, 8, and 24 hours. Variability between and within subjects was estimated by using analysis of variance for random effects. RESULTS: Intrasubject variability was highest for electric testing of the root and lowest for cold testing of the root across time. Of all subjects, 93% fell within 5 degrees C at all periods for the cold stimulation/moderate pain threshold. CONCLUSIONS: Calibrated cold stimulation of root areas appears to provide the most sensitive measure to assess therapeutic interventions to control cervical dental pain because of low intrasubject variability in untreated patients.

Neutrophil migration mechanisms, with an emphasis on the pulmonary vasculature.

Leukocyte trafficking into pulmonary tissue and airspaces is a critical component of the host defense response. Activation and migration of polymorphonuclear leukocytes (PMNs) into lungs also contribute to inflammatory tissue injury and remodeling of tissue architecture. There have been considerable advances in our understanding of the mechanisms that control PMN adhesion and transendothelial migration (TEM). Mechanisms of migration unique to the lungs have been described with regard to the profile of adhesion molecules, cytokines, and chemokines elicited during PMN emigration from blood vessels. This work reviews general mechanisms of TEM of PMNs and discusses the nature of PMN recruitment in several models of airway inflammation that illustrate how various stimuli elicit different responses. Pharmacologic manipulation of adhesive interactions between PMNs and endothelial cells is a current area of research aimed at developing pharmacologic agents to control inflammation during pulmonary and other inflammatory diseases. A summary of some of these agents and their actions is presented.

Neutrophil migration during endotoxemia.

Endotoxemia is marked by a global activation of inflammatory responses, which can lead to shock, multiple organ failure, and the suppression of immune and wound healing processes. Neutrophils (PMNs) play a central role in some of these responses by accumulating in tissues and releasing reactive oxygen species and proteases that injure host structures. This review focuses on altered PMN migratory responses that occur during endotoxemia and their consequences in the development of pulmonary infection. The inflammatory mediators that might be responsible for these altered responses are discussed. The oxidant potential of PMNs is increased after exposure to endotoxin both in vitro and during clinical and experimental endotoxemia. However, other functions such as chemotaxis and phagocytosis are often depressed in these same cells. Endotoxin exposure renders PMNs hyperadhesive to endothelium. The sum of these effects produces activated inflammatory cells that are incapable of leaving the vasculature. As such, the endotoxic PMN is more likely to promote tissue injury from within microvascular beds than to clear pathogens from extravascular sites. Moreover, the functional characteristics of endotoxic PMNs are similar to those observed during trauma, burn injury, sepsis, surgery, and other inflammatory conditions. Accordingly, several clinical conditions might have a common effector in the activated, yet migratorially dysfunctional, PMN. Direct effects of endotoxin on PMNs as well as effects of endogenous mediators released during endotoxemia are discussed. Understanding PMN behavior during endotoxemia may provide basic and critical insights that can be applied to a number of inflammatory scenarios.

Inhibition of pulmonary neutrophil trafficking during endotoxemia is dependent on the stimulus for migration.

In rat models of Gram-negative pneumonia, pulmonary emigration of neutrophils (polymorphonuclear leukocytes [PMNs]) is blocked when rats are made endotoxemic by an intravenous administration of endotoxin (lipopolysaccharide [LPS]). To test whether dysfunctional PMN migratory responses in the endotoxemic rat are specific for airway endotoxin, we gave rats intrapulmonary stimuli known to elicit different adhesion pathways for pulmonary PMN migration. Sprague-Dawley rats were treated intravenously with either saline or LPS and then instilled intratracheally with either sterile saline, LPS from Escherichia coli, interleukin (IL)-1, hydrochloric acid (HCl), zymosan-activated serum (ZAS), or lipoteichoic acid (LTA). Three hours later, accumulation of PMNs and protein in bronchoalveolar lavage fluid (BALF) were assessed. BALF PMN accumulation in response to intratracheal treatment with LPS (100%), IL-1 (100%), ZAS (40%), and LTA (58%) was inhibited by endotoxemia. In rats given intratracheal HCl, BALF PMN numbers were unaffected by intravenous LPS. The pattern of inhibition of migration suggests that intravenous LPS only inhibits migration in response to stimuli for which migration is CD18-dependent. In contrast to PMN migration, BALF protein accumulation was inhibited by intravenous LPS only when IL-1 or LPS was used as the intratracheal stimulus. To characterize further the differential responses to the various airway stimuli, the appearance in BALF of tumor necrosis factor-alpha (TNF-alpha) and the PMN chemokine macrophage inflammatory protein (MIP)-2 was measured. Accumulation of PMNs in BALF correlated with the BALF concentrations of MIP-2 (r = 0.846, P < 0.05) and TNF (r = 0.911; P < 0.05). The ability of intravenous LPS to inhibit pulmonary PMN migration correlated weakly with MIP-2 (r = 0.659; P < 0.05) and with TNF (r = 0.413; P > 0.05) concentrations in BALF. However, this correlation was strengthened for TNF (r = 0.752; P < 0.05) when data from IL-1-treated animals were excluded. Thus, the presence in BALF of inflammatory mediators that are known to promote CD18-mediated migration correlates with endotoxemia-related inhibition of PMN migration. Furthermore, the pattern of inhibition of pulmonary PMN migration during endotoxemia is consistent with the CD18 requirement of each migratory stimulus.

Interaction of rat Pneumocystis carinii and rat alveolar epithelial cells in vitro.

During Pneumocystis carinii pneumonia, P. carinii trophic forms adhere tightly to type I alveolar epithelial cells (AECs). However, the manner in which the interaction between P. carinii organisms and AECs results in clinical pneumonia has not been explored. To investigate this interaction in vitro, we established a culture system using rat P. carinii and primary cultures of rat AECs. We hypothesized that binding of P. carinii to AECs would alter the metabolic, structural, and barrier functions of confluent AECs. Using fluorescently labeled P. carinii, we demonstrated that P. carinii bound to AECs in a dose-dependent manner. During P. carinii-AEC interaction, both the AECs and the P. carinii organisms remained metabolically active. Immunofluorescent staining demonstrated that AEC expression of the junctional proteins E-cadherin and occludin and the structural protein cytokeratin 8 were unaffected by P. carinii binding. To evaluate the effect of P. carinii on AEC barrier function, transepithelial resistance across AEC monolayers was measured during interaction with organisms. Culture with P. carinii did not result in loss of AEC barrier function but in fact increased AEC transepithelial resistance in a dose- and time-dependent manner. We conclude that the direct interaction of P. carinii with AECs does not disrupt AEC metabolic, structural, or barrier function. Therefore, we speculate that additional inflammatory cells and/or their signals are required to induce the epithelial derangements characteristic of P. carinii pneumonia.

Continuous amperometric monitoring of glucose in a brittle diabetic chimpanzee with a miniature subcutaneous electrode.

The performance of an amperometric biosensor, consisting of a subcutaneously implanted miniature (0.29 mm diameter, 5 x 10(-4) cm2 mass transporting area), 90 s 10-90% rise/decay time glucose electrode, and an on-the-skin electrocardiogram Ag/AgCl electrode was tested in an unconstrained, naturally diabetic, brittle, type I, insulin-dependent chimpanzee. The chimpanzee was trained to wear on her wrist a small electronic package and to present her heel for capillary blood samples. In five sets of measurements, averaging 5 h each, 82 capillary blood samples were assayed, their concentrations ranging from 35 to 400 mg/dl. The current readings were translated to blood glucose concentration by assaying, at t = 1 h, one blood sample for each implanted sensor. The rms error in the correlation between the sensor-measured glucose concentration and that in capillary blood was 17.2%, 4.9% above the intrinsic 12.3% rms error of the Accu-Chek II reference, through which the illness of the chimpanzee was routinely managed. Linear regression analysis of the data points taken at t>1 h yielded the relationship (Accu-Chek) = 0. 98 x (implanted sensor) + 4.2 mg/dl, r2 = 0.94. The capillary blood and the subcutaneous glucose concentrations were statistically indistinguishable when the rate of change was less than 1 mg/(dl. min). However, when the rate of decline exceeded 1.8 mg/(dl.min) after insulin injection, the subcutaneous glucose concentration was transiently higher.

Right ventricular end-diastolic volume as a predictor of the hemodynamic response to a fluid challenge.

OBJECTIVE: To compare thermodilution right ventricular end-diastolic volume index (RVEDVI) and pulmonary artery occlusion pressure (Ppao) as predictors of the hemodynamic response to a fluid challenge. DESIGN: Prospective cohort study. SETTING: Medical ICU of a university-affiliated county hospital and medical-surgical ICU of a community hospital. PATIENTS: Twenty-five critically ill patients who had one or more clinical conditions that suggested the possibility of inadequate preload. INTERVENTIONS: Thirty-six fluid challenges. Fluid (saline or colloid) was administered rapidly until the Ppao rose by at least 3 mm Hg. When a patient underwent more than one fluid challenge, these were given on separate days and for different clinical indications. MEASUREMENTS AND RESULTS: Responders (n=20; > or = 10% increase in stroke volume [SV]) and nonresponders (n=16; <10% increase in SV) differed with respect to baseline Ppao (10.0+/-3.4 vs 14.2+/-3.6 mm Hg; p=0.001), but not with respect to baseline RVEDVI (105+/-31 vs 119+/-33 mL/m2; p=0.22). There was a moderate correlation between RVEDVI and fluid-induced change in SV (r=0.44); the relationship between Ppao and change in SV was stronger (r=0.58). A positive response to fluid was observed in 4 of 9 cases in which RVEDVI exceeded 138 mL/m2, a threshold value that has been suggested to reliably predict a poor response to fluid. CONCLUSION: RVEDVI was not a reliable predictor of the response to fluid. As a predictor of fluid responsiveness, Ppao was superior to RVEDVI. In an individual patient, adequacy of preload is best assessed by an empiric fluid challenge.

Acute group G streptococcal myositis associated with streptococcal toxic shock syndrome: case report and review.

The group G streptococcus (GGS) is a rare cause of deep soft-tissue infection. We report that we believe is the first case of acute diffuse GGS myositis in association with toxic shock. Although the causative organism did not contain the genes for group A streptococcal pyrogenic exotoxins (SPEs) or make SPEs, the organism produced at least one new toxin that shared the biologic properties of the SPEs.

Nitric oxide is not involved in hepatocyte killing by neutrophils activated by N-formyl-methionyl-leucylphenylalanine or phorbol myristate acetate in vitro.

Polymorphonuclear leukocytes (PMNS) have been implicated as cellular mediators of hepatic injury in models of inflammation in vivo. In vitro, hepatocyte killing by activated PMNs is mediated in part by proteases, but the role of nitric oxide is unknown. NO is produced by PMNs and hepatocytes and can act either to damage or protect in various models of toxicity. Therefore, we tested the hypothesis that NO is important in PMN-mediated hepatocyte killing in vitro. Freshly isolated hepatocytes from rat liver and PMNs elicited from rat peritoneum were cultured together or alone for 16 hours. Both cell types spontaneously released NO, estimated as its stable breakdown product, nitrite. Accumulation of nitrite in medium from hepatocyte cultures was augmented threefold by incubation with L-arginine and was completely inhibited by treatment with the nitric oxide synthase (NOS) inhibitor NG-methyl-L-arginine (NMA). Nitrite release in PMN cultures was unaffected by L-arginine addition and only partially inhibited by NMA. In PMN:hepatocyte cocultures (10:1), accumulation of nitrite was additive relative to cells cultured separately. Incubation with NMA blocked nitrite production completely in cocultures, whereas L-arginine caused a two-fold increase in nitrite. Addition of PMN stimulants, N-formyl-methionyl-leucyl-phenylalanine (FMLP), or phorbol myristate acetate (PMA), caused increased release of alanine aminotransferase (ALT) activity into medium from hepatocytes cultured with PMNs but not from hepatocytes cultured alone; this indicated that injury to hepatocytes was due to activated PMNS. However, neither FMLP nor PMA significantly altered nitrite release from cocultures. Despite the alterations in NO production induced by addition of NMA or L-arginine, neither agent altered the release of ALT from hepatocytes in coculture with activated PMNs. Thus, PMNs and hepatocytes provided NO in vitro, but neither suppression nor elevation of NO production affected PMN-mediated hepatocyte killing. Accordingly, NO is not involved in the mechanisms by which FMLP-or PMA-stimulated PMNs mediate hepatocyte injury in vitro.

Kinetics of glucose delivery to subcutaneous tissue in rats measured with 0.3-mm amperometric microsensors.

The time between intravenous injection of a glucose bolus and the time the glucose concentration peaked in the subcutaneous tissue was measured in pentobarbital-anesthetized rats with implanted 290-microns-diameter amperometric sensors. Boluses of 100, 200, and 400 mg/kg body wt were injected. The glucose concentration in the jugular vein was monitored by frequent withdrawal and analysis of samples. The glucose concentration in the subcutaneous tissue was continuously monitored with the sensors. The times required for the subcutaneously implanted sensor to reach its maximum current, corrected for sensor response times, were 7.5 +/- 3.9, 9.8 +/- 5.5, and 10.0 +/- 4.4 min for the smallest to the largest dose, respectively. The shorter delay in response to the smallest dose was statistically significant (P < 0.03). The results were consistent with dilution of the bolus in the cardiovascular system and transport of glucose by both diffusion and facilitated transport via a saturable mediator. An understanding of the differences in the dynamics of venous vs. subcutaneous response to a glucose dose is important in developing algorithms for the control of blood glucose based on a subcutaneous measurement.

Toxicity of mitomycin C toward cultured pulmonary artery endothelium.

Mitomycin C (MMC) is a bifunctional alkylating agent used in cancer chemotherapy. MMC therapy occasionally results in pulmonary vascular injury, including alterations in endothelial cells. Reactive metabolites of MMC can cross-link DNA, and its cytotoxicity has been attributed in part to this capacity, but effects in vascular cells have not been explored extensively. Accordingly, the direct effects of MMC on cultured porcine pulmonary artery endothelial cells (PECs) were examined. A single administration of MMC (0-10 microM) to PEC monolayers resulted in concentration-dependent cytolytic injury that was delayed in onset and progressive in nature. Cells treated at subconfluent densities were inhibited in their ability to proliferate. MMC treatment resulted in DNA cross-linking at concentrations (0.01-1 microM) that inhibited cell proliferation but caused only limited overt cytotoxicity, supporting an association between DNA cross-linking and impairment of cell division. This pattern of PEC injury is reminiscent of that seen after treatment with another pneumotoxic, bifunctional alkylating agent, monocrotaline pyrrole. The similarity of the endothelial cell response to different bifunctional alkylating agents suggests that DNA cross-linking may inhibit cell proliferation and thereby limit the repair capacity of endothelial monolayers. This might contribute to the progressive pulmonary vascular injury that occurs after administration of certain DNA cross-linking agents in vivo.

Lactate dehydrogenase activity and isozyme patterns in tissues and bronchoalveolar lavage fluid from rats treated with monocrotaline pyrrole.

Monocrotaline pyrrole (MCTP), a putative, toxic metabolite of monocrotaline, induces delayed and progressive lung injury, vascular remodeling, and pulmonary hypertension in rats. The lung injury is characterized by increased wet lung-to-body weight ratio followed by increases in lactate dehydrogenase (LDH) activity and protein concentration in the cell-free bronchoalveolar lavage fluid (BALF) and increased cellularity of BALF. We evaluated total LDH activity and isozyme patterns in the tissues, cell lysates, sera and cell-free BALF of rats after treatment with MCTP to determine the source of increased LDH activity. Male Sprague-Dawley rats were given a single injection of MCTP (3.5 mg/kg) or an equal volume of the N,N-dimethylformamide (DMF) vehicle in the tail vein on Day 0. Rats were killed at 4, 8, or 14 days after toxicant administration, and several markers of lung injury, LDH activity, and isozyme patterns of various tissues, cells, and body fluids were determined. At 8 and 14 days, the lungs from MCTP-treated rats had multifocal, irregularly shaped lesions of hemorrhage and consolidation. At Day 14 only, the hearts of MCTP-treated rats appeared enlarged and there was right cardioventricular hypertrophy. Rats treated with MCTP had no macroscopic lesions in kidneys, liver, or skeletal muscle. Compared to controls, MCTP-treated animals had no change in total LDH activity or isozyme patterns of samples of lungs, heart, skeletal muscle, liver, kidneys, or erythrocyte lysates. Changes in LDH activity in the cell-free BALF and BALF cell pellet from rats treated with MCTP were characterized by increases in isozymes LDH4 and LDH5 and an elevated LDH4/LDH5 ratio in the BALF only. Our results suggest the most probable source of the increased LDH activity in cell-free BALF of MCTP-treated rats originates from the lung tissue and is consistent with a contribution from the pulmonary vascular endothelium, a source rich in LDH4. A combination of plasma, macrophages, and neutrophils in the pulmonary tissue may also have made minor contributions to the increase in cell-free BALF LDH activity, particularly to the activity of LDH5.