Cod Liver Oil, but Not Retinoic Acid, Treatment Restores Bone Thickness in a Vitamin A-Deficient Rat.

Vitamin A plays a prominent role for maintaining optimal bone status, but its impact upon the bone in response to vitamin A deficiency is not well defined. The purpose of this study was to evaluate how replenishing vitamin A by either whole food cod liver oil (COD) or the active metabolite of vitamin A, retinoic acid (RA), altered bone thickness of vitamin A-deficient (VAD) rats. Weanling rats were administered a control diet (CTRL) or VAD diet for 9 weeks. This was followed by four weeks of treatment in which the VAD group was divided into the following 4 subgroups: (1) VAD (9 weeks)-VAD (4 weeks); (2) VAD-CTRL; (3) VAD-COD; and (4) VAD-RA. Compared to controls, VAD rats had thicker bones which showed marked dysplasia. VAD-rats treated with COD produced a thinner bone that was not significantly different from that of untreated rats. In contrast, RA did not significantly change the thicker bone, and also had significantly greater periosteal and endosteal osteoblast numbers compared to VAD-COD. Active osteoclasts were not detected in VAD rats, nor during the treatment period. These findings suggest that the abnormal bone thickness in VAD rats appears to be more effectively restored to bone thickness of untreated control rats when treated with COD.

Fat embolism sensitizes rats to a "second hit" with lipopolysaccharide: An animal model of pulmonary fibrosis.

BACKGROUND: Pulmonary fat embolism (FE) in patients after major bone fracture and other trauma may lead to acute respiratory distress, but few clinical evidence of lung injury remains, and there is a dearth of histopathologic information after the initial recovery. We recently reported histologic changes in the lungs of a patient who died after cesarian delivery, which were similar to a rat model of FE. In this model, we found that despite an apparent full recovery, modest fibrotic damage persisted up to 6 weeks. We tested whether at that time, an additional insult could exacerbate the effects. METHODS: Triolein (0.2 mL intravenously administered) was given to 18 rats and saline to 18 controls. Six weeks later, each group received (intraperitoneal) lipopolysaccharide (LPS, 3 mg/kg; n = 9) or saline (n = 9). At necropsy 48 hours later, lungs and organs were harvested for study. Lung parenchymal, vascular, and bronchial damage was scored by two pathologists and by Image J analysis. RESULTS: Animals given LPS after triolein showed reduced pulmonary arterial medial diameters compared with those that received LPS alone (p < 0.04). Lung small arterial patency (lumen) was reduced after triolein and even more after combined LPS and triolein (p = 0.018). Triolein increased fibrotic markers (trichrome and smooth muscle actin staining), and this was more severe after LPS. At 6 weeks, fat droplets remained in the lungs, localizing to the subpleural septa. These were smaller and more widespread after LPS. CONCLUSION: This report describes an animal model to study exacerbation of lung histopathology induced by FE using a known pulmonary toxicant, LPS (a "second hit"). Vascular and fibrotic lung damage was more severe when LPS was given to rats 6 weeks after triolein compared with LPS alone. FE rendered the lungs extra sensitive to a second hit long after apparent clinical recovery. This experimental model of fat embolism provides useful informations for the treatment of patients suffering for similar conditions.

Role of anthocyanin-enriched purple-fleshed sweet potato p40 in colorectal cancer prevention.

SCOPE: Anthocyanins, the natural pigments in plant foods, have been associated with cancer prevention. However, the content of anthocyanins in staple foods is typically low and the mechanisms by which they exert anticancer activity is not yet fully defined. METHODS AND RESULTS: We selected an anthocyanin-enriched purple-fleshed sweet potato clone, P40, and investigated its potential anticancer effect in both in vitro cell culture and in vivo animal model. In addition to a high level of total phenolics and antioxidant capacity, P40 possesses a high content of anthocyanins at 7.5 mg/g dry matter. Treatment of human colonic SW480 cancer cells with P40 anthocyanin extracts at 0-40 µM of peonidin-3-glucoside equivalent resulted in a dose-dependent decrease in cell number due to cytostatic arrest of cell cycle at G1 phase but not cytotoxicity. Furthermore, dietary P40 at 10-30% significantly suppressed azoxymethane-induced formation of aberrant crypt foci in the colons of CF-1 mice in conjunction with, at least in part, a lesser proliferative PCNA and a greater apoptotic caspase-3 expression in the colon mucosal epithelial cells. CONCLUSION: These observations, coupled with both in vitro and in vivo studies reported here, suggest anthocyanin-enriched sweet potato P40 may protect against colorectal cancer by inducing cell-cycle arrest, antiproliferative, and apoptotic mechanisms.

Dietary Flaxseed Oil Protects against Bleomycin-Induced Pulmonary Fibrosis in Rats.

Bleomycin, a widely used antineoplastic agent, has been associated with severe pulmonary toxicity, primarily fibrosis. Previous work has shown a reduction in bleomycin-induced lung pathology by long-chain omega-3 fatty acids. Treatment by short-chain omega-3 fatty acids, α-linolenic acid, found in dietary flaxseed oil may also reduce lung fibrosis, as previously evidenced in the kidney. To test this hypothesis, 72 rats were divided between diets receiving either 15% (w/w) flaxseed oil or 15% (w/w) corn oil (control). These groups were further divided to receive either bleomycin or vehicle (saline) via an oropharyngeal delivery, rather than the traditional intratracheal instillation. Lungs were harvested at 2, 7, and 21 days after bleomycin or saline treatment. Animals receiving flaxseed oil showed a delay in edema formation (P = 0.025) and a decrease in inflammatory cell infiltrate and vasculitis (P = 0.04 and 0.007, resp.). At days 7 and 21, bleomycin produced a reduction in pulmonary arterial lumen patency (P = 0.01), but not in rats that were treated with flaxseed oil. Bleomycin-treated rats receiving flaxseed oil had reduced pulmonary septal thickness (P = 0.01), signifying decreased fibrosis. Dietary flaxseed oil may prove beneficial against the side effects of this highly effective chemotherapeutic agent and its known toxic effects on the lung.

Effect of exendin (exenatide)--GLP 1 receptor agonist on the thyroid and parathyroid gland in a rat model.

Exenatide or Exendin-4 is a 39-amino acid agonist of the glucagon like peptide (GLP-1) receptor approved for the adjunctive treatment for type 2 diabetes. Recent reports suggest that GLP-1 agonists may also have distant effects including C-cell thyroid hyperplasia. The aim of this study was to evaluate the effect of exendin-4 on the thyroid and parathyroid cells in a rat model. Rat thyroids were stained for calcitonin, H&E and for carcinoembryonic antigen (CEA). Thyroid C-cell hyperplasia was graded on H&E stained slides using cell size and secretory granule numbers, morphological features of the parathyroid glands and the serum calcium concentrations of the rats were also evaluated. Counts of stained cells/high power field and intensity of staining were recorded by two pathologists. Data were analyzed by ANOVA/post-tests. C cell hypertrophy was elevated in exenatide-treated vs. untreated animals (22.5 ± 8.7 vs. 10.5 ± 2.7 cells/HPF). CEA staining failed to show effects by exendin. Calcitonin staining was significantly elevated in exenatide treated controls (P<0.001). Parathyroid glands were histologically normal in both groups, and serum calcium levels were within normal range in all animals. In summary, exenatide was associated with C cell hyperplasia and increased calcitonin staining of thyroids, but was unrelated to CEA levels. These data raise important concerns about the effects of exenatide which, given its wide clinical use, should be clarified with urgency.

Persistent and progressive pulmonary fibrotic changes in a model of fat embolism.

BACKGROUND: Fat embolism (FE) after trauma and some orthopedic procedures is known to cause acute lung injury, including acute respiratory distress syndrome. However, its potential long-term effects on the lung are unknown. A previous study using a rat model of FE found significant histopathologic changes in the lungs after intravenous injection of triolein for up to 11 days. This study detailed the persistence of the lung damage and investigated the input of the renin-angiotensin system in its pathology. METHODS: Unanesthetized rats were injected via the tail vein with 0.2 mL saline or triolein. After euthanasia, at 3 weeks or 6 weeks, lung sections were stained to highlight cellular structure, presence of collagen and fat, or immunolabeled for smooth muscle actin or angiotensin peptides. RESULTS: At 3 weeks or 6 weeks after triolein injection, there was no dilatation of the heart or inferior vena cava, no congestion of the liver or spleen, no adventitial edema, nor was fluid present in alveoli or pleural cavity as reported in animals at earlier time points. Persisting pathology included reduced lumen patency, thickening of the media of small arteries and arterioles, and vascular and septal inflammation. Although the fat content of the lung decreased from week 3 to week 6, there was a progressive increase in collagen, smooth muscle actin, and angiotensin peptides. CONCLUSIONS: This model extends the effect of FE on pulmonary pathology to 6 weeks, revealing persistent vasculitis, septal inflammation, and progressive fibrotic changes which are associated with increased presence of angiotensin peptides.

Mitigating effects of captopril and losartan on lung histopathology in a rat model of fat embolism.

BACKGROUND: Fat embolization (FE) is an often overlooked and poorly understood complication of skeletal trauma and some orthopedic procedures. Fat embolism can lead to major pulmonary damage associated with fat embolism syndrome (FES). METHODS: A model of FE in unanesthetized rats, using intravenous injection of the neutral fat triolein, was used to study the potential therapeutic effect on lung histopathology of altering the production of, or response to, endogenous angiotensin (Ang) II. Either captopril, an Ang I converting enzyme inhibitor, or losartan, an Ang II type 1 receptor blocker, was injected 1 hour after FE by triolein injection. After euthanasia at 48 hours, histopathologic evaluation was used to compare the drug-treated animals with control animals that received only triolein. RESULTS: Histology of the lungs of rats treated only with triolein revealed severe, diffuse pathology. Alveolar septa showed severe, diffuse inflammation. Bronchial lumina showed severe mucosal epithelial loss. The media of the pulmonary small arteries and arterioles was thicker, and the lumen patency was reduced 60% to 70%. Trichrome staining confirmed the abundant presence of collagen in the media and adventitia, as well as collagen infiltrating the bronchial musculature. Both captopril and losartan treatments reduced the inflammatory, vasoconstrictor, and profibrotic effects present at 48 hours (p<0.001). With treatment, the vascular lumen remained patent, and the fat droplets were reduced in size and number. There was a reduction in the number of infiltrating leukocytes, macrophages, myofibroblasts, and eosinophils, along with a significant decrease in hemorrhage and collagen deposition (p<0.001). Pathologic changes in bronchial epithelium were also diminished. CONCLUSIONS: The results suggest that the use of drugs that act on the renin-Ang system might provide an effective and targeted therapy for fat embolism syndrome.

Treatment with omega-3 fatty acids but not exendin-4 improves hepatic steatosis.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is the most common form of chronic liver disease in the Western world. The aim of this study was to evaluate the biochemical and histological effects of omega-3 fatty acid and exendin-4 treatment on NAFLD in an animal model. METHODS: Sixty-three 8-week-old outbred Sprague-Dawley male rats were used for this study. Three animals were used as procedure controls, and 30 rats were fed a methionine and choline deficient (MCD) diet and 30 were fed a regular chow diet. In each group of 30 animals, 10 served as controls, 10 received exendin-4, and 10 received omega-3 fatty acids. After 75 days of treatment, the animals were euthanized, the tissues and serum were harvested, and the livers were formalin-fixed for histology. RESULTS: The MCD diet was exceptionally efficient at producing fatty livers. The MCD control animals had a liver steatosis score of 38+/-6.7 (of 50 possible); treatment with exendin-4 was not associated with a significant reduction of steatosis (44+/-5.16, P=0.07) and the omega-3 fatty acid treatment was associated with a significant decrease in the liver steatosis score (15.6+/-13.46, P<0.001) compared with both the controls and the exendin-4 groups. The omega-3 fatty acid treatment increased serum aspartate aminotransferase significantly, whereas exendin-4 had no effect. CONCLUSION: In an animal model of NAFLD, the omega-3 fatty acid therapy was associated with significant improvement in hepatic steatosis compared with exendin-4. These data suggest that omega-3 fatty acid supplements may have a potential therapeutic role in patients with NAFLD.

Weight Loss via exercise with controlled dietary intake may affect phospholipid profile for cancer prevention in murine skin tissues.

Exercise has been linked to a reduced cancer risk in animal models. However, the underlying mechanisms are unclear. This study assessed the effect of exercise with dietary consideration on the phospholipid profile in 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced mouse skin tissues. CD-1 mice were randomly assigned to one of the three groups: ad libitum-fed sedentary control; ad libitum-fed treadmill exercise at 13.4 m/min for 60 min/d, 5 d/wk (Ex+AL); and treadmill-exercised but pair-fed with the same amount as the control (Ex+PF). After 14 weeks, Ex+PF but not Ex+AL mice showed approximately 25% decrease in both body weight and body fat when compared with the controls. Of the total 338 phospholipids determined by electrospray ionization-tandem mass spectrometry, 57 were significantly changed, and 25 species could distinguish effects of exercise and diet treatments in a stepwise discriminant analysis. A 36% to 75% decrease of phosphatidylinositol (PI) levels in Ex+PF mice occurred along with a significant reduction of PI 3-kinase in TPA-induced skin epidermis, as measured by both Western blotting and immunohistochemistry. In addition, approximately 2-fold increase of the long-chain polyunsaturated fatty acids, docosahexaenoic and docosapentaenoic acids, in phosphatidylcholines, phosphatidylethanolamines, and lysophosphatidylethanolamines was observed in the Ex+PF group. Microarray analysis indicated that the expression of fatty acid elongase-1 increased. Taken together, these data indicate that exercise with controlled dietary intake, but not exercise alone, significantly reduced body weight and body fat as well as modified the phospholipid profile, which may contribute to cancer prevention by reducing TPA-induced PI 3-kinase and by enhancing omega-3 fatty acid elongation.

Fat embolism: evolution of histopathological changes in the rat lung.

The pathophysiology of Fat Embolism Syndrome (FES) is poorly understood and subject to some controversy. Evaluation of the evolution of histological changes in the lungs of patients with FES is impractical. The current theories of FES were established through acute clinical observations and acute animal experiments, but sequential changes in the histology of lungs over a prolonged period have not been made. The progressive effects of fat embolization of the lungs were examined in a rat model over a period of 11 days. Triolein, a major bone marrow fat, was administered to conscious Sprague-Dawley rats via the caudal vein. Rats were euthanized at 24, 48, 96 h, and 11 days, but some died within a few hours. Histomorphometric evaluations of lung tissue were made, including stains for fat, collagen, and smooth muscle actin. Arterial and arteriolar patency decreased progressively up to 96 h, but returned toward normal after 11 days. A striking finding was the very early presence of inflammation and fibrosis after only several hours, persisting up to 11 days. The results of this study provide evidence of both very early and prolonged changes due to fat embolization.

Expression of multidrug resistance associated protein 5 (MRP5) on cornea and its role in drug efflux.

PURPOSE: The purpose of this manuscript is to investigate the presence of nucleoside/nucleotide efflux transporter in cornea and to evaluate the role in ocular drug efflux. METHODS: RT-PCR, immunoprecipitation followed by Western blot analysis and immunostaining were employed to establish molecular presence of multidrug resistance associated protein 5 (MRP5) on cornea. Corneal efflux by MRP5 was studied with bis(POM)-PMEA and acyclovir using rabbit and human corneal epithelial cells along with MRP5 over expressing cells (MDCKII-MRP5). Ex vivo studies using excised rabbit cornea and in vivo ocular microdialysis in male New Zealand white rabbits were used to further evaluate the role of MRP5 in conferring ocular drug resistance. RESULTS: RT-PCR confirms the expression of MRP5 in both rabbit and human corneal epithelial cells along with MDCKII-MRP5 cells. Immunoprecipitation followed by Western blot analysis using a rat (M511-54) monoclonal antibody that reacts with human epitope confirms the expression of MRP5 protein in human corneal epithelial cells and MDCKII-MRP5 cells. Immunostaining performed on human cornea indicates the localization of this efflux pump on both epithelium and endothelium. Efflux studies reveal that depletion of ATP decreased PMEA efflux significantly. MRP5 inhibitors also diminished PMEA and acyclovir efflux. However, depletion of glutathione did not alter efflux. MDR1 and MRP2 did not contribute to PMEA efflux. However, MRP2 is involved in acyclovir efflux while MDR1 do not participate in this process. TLC/autoradiography suggested the conversion of bis(POM)-PMEA to PMEA in rabbit and human corneal epithelial cells. Two well known antiglaucoma drugs, bimatoprost and latanoprost were rapidly effluxed by MRP5. Ex vivo study on intact rabbit corneas demonstrated accumulation of PMEA in cornea in the presence of ATP-depleting medium. In vivo ocular pharmacokinetics also revealed a significant increase in maximum aqueous humor concentration (C(max)) and area under the aqueous humor time curve (AUC) of acyclovir in the presence of MK-571, a specific MRP inhibitor. CONCLUSIONS: Taken together immunolocalization on human cornea, in vitro efflux in human, rabbit corneal and MRP5 over expressing cells, ex vivo and in vivo studies in intact rabbit cornea suggest that MRP5 on cornea can significantly lower the permeability of antiviral and glaucoma drugs. These findings may be valuable in developing formulation strategies to optimize ocular bioavailability of topically administered ocular agents.

Metabolic and biochemical responses of the healthy human lung to nonthoracic surgery.

The objective of this study was to evaluate and assign numbers to biochemical or cellular entities in lung-healthy patients that change immediately postsurgery compared with the same parameters immediately presurgery, with the hypothesis that biochemical markers with significant change could be the basis of tests to predict postoperative respiratory complications. Thirty lung-healthy adults who were to undergo elective surgical procedures requiring general anesthesia participated. The population included sequential persons that met inclusion criteria and gave consent. At intubation and before surgery, a bonchoalveolar lavage (BAL) was performed. Before extubation but after completion of surgical procedures, a second 100-ml BAL was performed in the contralateral lung. Serum from both time periods was also collected. Total cell counts were elevated postsurgery in smokers and subjects claiming childhood but not current asthma, who also showed increased postsurgical BAL IL-1 but not increased TNFalpha. LDH and its isoenzymes, measured in both BAL and serum, showed no correlation with time on surgical ventilation, average FiO2, or average peak pressure during surgical ventilation. BAL LDH isoenzyme 4 showed a significant elevation pattern pre-to-post surgery when the entire subject population was considered irrespective of surgery type or time on ventilation. Presurgery versus postsurgery variation was best measured in BAL rather than in serum. The pulmonary, biochemical, and cellular parameters measured in the pre- and postsurgical BALs of lung-healthy subjects undergoing nonthoracic surgery show subtle modulations of pulmonary defense markers, defined by significantly increased proinflammatory cytokines and cell counts postsurgery compared to the same patient presurgery.

Effects of dietary calorie restriction or exercise on the PI3K and Ras signaling pathways in the skin of mice.

Weight control by exercise and dietary calorie restriction (DCR) has been associated with reduced cancer risk, but the underlying mechanisms are not well understood. This study was designed to compare the effects of weight loss by increasing physical activity or decreasing calorie intake on tumor promoter-induced Ras-MAPK and PI3K-Akt pathways. SENCAR mice were randomly assigned to one of the following five groups: ad libitum-fed sedentary control, ad libitum-fed exercise (AL+Exe), exercise but pair-fed at the amount as controls (PF+Exe), 20% DCR, and 20% DCR plus exercise (DCR+Exe). After 10 weeks, body weight and body fat significantly decreased in the groups of DCR, DCR+Exe, and PF+Exe when compared with the controls. AL+Exe did not induce weight loss due to, at least in part, increased food intake. Plasma IGF-1 levels reduced significantly in DCR and DCR+Exe but not PF+Exe. The protein H-Ras and activated Ras-GTP significantly decreased in TPA-induced skin tissues of DCR-fed mice but not exercised mice. PI3K protein, phosphoserine Akt, and p42/p44-MAPK were reduced, however, in both DCR and PF+Exe groups. Immunohistochemistry demonstrated that the significantly reduced H-Ras occurred in subcutaneous fat cells, while the reduced PI3K and PCNA took place only in the epidermis. Plasma leptin decreased in PF+Exe, DCR, and DCR+Exe, while the caspase-3 activity increased in DCR+Exe only. Genomic microarray analysis further indicated that the expression of 34 genes relevant to PI3K and 31 genes to the MAPK pathway were significantly regulated by either DCR or PF+Exe treatments. The reduced PI3K in PF+Exe mice was partially reversed by IGF-1 treatment. The overall results of this study demonstrated that DCR abrogated both Ras and PI3K signaling, which might inhibit TPA-induced proliferation and anti-apoptosis. Selective inhibition of PI3K by PF+Exe but not AL+Exe seems more attributable to the magnitude of the caloric deficit and/or body fat loss than diet versus exercise comparison.

Effect of an angiotensin II receptor blocker and two angiotensin converting enzyme inhibitors on transforming growth factor-beta (TGF-beta) and alpha-actomyosin (alpha SMA), important mediators of radiation-induced pneumopathy and lung fibrosis.

Progressive, irreversible fibrosis is one of the most clinically significant consequences of ionizing radiation on normal tissue. When applied to lungs, it leads to a complication described as idiopathic pneumonia syndrome (IPS) and eventually to organ fibrosis. For its high mortality, the condition precludes treatment with high doses of radiation. There is widespread interest to understand the pathogenetic mechanisms of IPS and to find drugs effective in the prevention of its development. This report summarizes our experience with the protective effects of L 158,809, an angiotensin II (ANG II) receptor blocker, and two angiotensin converting enzyme (ACE) inhibitors in the development of IPS and the role of transforming growth factor beta (TGF-beta) and of alpha-actomyosin (alpha SMA) in pathogenesis of radiation induced pulmonary fibrosis in an experimental model of bone marrow transplant (BMT). Male WAG/Riji/MCV rats received total body irradiation and a regimen of cyclophosphamide (CTX) in preparation for bone marrow transplant. While one group of animals remained untreated, the remainders were subdivided into three groups, each of them receiving either the ANG II receptor blocker or one of the two ACE inhibitors (Captopril or Enalapril). Each of the three drugs was administered orally from 11 days before the transplant up to 56 days post transplant. At sacrifice time the irradiated rats receiving only CTX showed a chronic pneumonitis with septal fibrosis and vasculitis affecting, in particular, small caliber pulmonary arteries and arterioles. Their lung content of hydroxyproline was also markedly elevated in association with the lung concentrations of thromboxane (TXA2) and prostaglandin (PGI(2)), (two markers of pulmonary endothelial damage). A significant increase of alpha actomyosin staining was observed in vessels, septa and macrophages of the same animals which also overexpressed TGF-beta. When L 158,809, Captopril and Enalapril were added to the radiation and cytoxan treatment, a significant amelioration of the histological damage as well as the overexpression of alpha SMA was observed. Lung concentrations of hydroxyproline, PGI(2), TXA2 and TGF-beta were also observed in these animals so that the values of these compounds were closer to those measured in untreated control rats than to their irradiated and cytoxan treated counterparts. Angiotensin II plays an important role in the regulation of TGF-beta and alpha SMA, two proteins involved in the pathogenesis of pulmonary fibrosis. The finding that ACE inhibitors or ANG II receptor blockers protect the lungs from radiation induced pneumonitis and fibrosis reaffirms the role that ANG II plays in this inflammatory process and suggests an additional indication of treatment of this condition, thus opening a new potential pharmacologic use of these drugs.

Effects on cytokines and histology by treatment with the ACE inhibitor captopril and the antioxidant retinoic acid in the monocrotaline model of experimentally induced lung fibrosis.

Monocrotaline (MCT), a pyrrolizidine alkaloid extracted from the shrub Crotalaria spectabilis, induces in the lungs of many mammalian species severe hypertension and fibrosis. Previous work with MCT-induced lung disease in rats has shown that some of the steps to progressive fibrosis can be interrupted or decreased by intervention with retinoic acid (RA) or with the angiotensin converting enzyme inhibitor, captopril. This report emphasizes the pathology and cytokines present in lungs of rats in the MCT model of hypertension and fibrosis in 8 treatment groups, six per group: (1) controls, not treated; (2) captopril; (3) RA; (4) combined captopril and RA. Groups 5-8 replicated groups 1-4 and also received MCT subcutaneously. Tissues were harvested at 28 days for histopathology and measurement of cytokines TGFbeta, TNFalpha, interleukin 6, and IFN_. TGFbeta was depressed at 28 days by MCT, an effect reversed by a combination of captopril and RA. RA influences production of an important Th1 cytokine, IFN_, and demonstrated the greatest limitation of MCT-induced TNFalpha. The MCT-induced lung pathology of vasculitis, interstitial pneumonia and fibrosis was limited by captopril. Smooth muscle actin was overexpressed in MCT treated animals receiving RA, an effect reduced with captopril. Overall, the study confirmed the existence of a protective effect for both captopril and RA from MCT-induced lung damage at 30 days. No synergistic or antagonistic activity was observed when the two drugs were administered together. Each of the drugs exerts different and particular effects on serum and tissue levels of various cytokines, suggesting that each drug is efficient at different points of attack in the control of lung fibrosis.

Phthalate-Induced Liver Protection against Deleterious Effects of the Th1 Response: A Potentially Serious Health Hazard.

Infection with Mycobacterium tuberculosis (TB) induces pulmonary immunopathology mediated by classical Th1 type of acquired immunity with hepatic involvement in up to 80% of disseminated cases. Since PPAR agonists cause immune responses characterized by a decrease in the secretion of Th1 cytokines, we investigated the impact of activating these receptors on hepatic pathology associated with a well-characterized model of Th1-type pulmonary response. Male Fischer 344 rats were either maintained on a drug-free diet (groups I and II), or a diet containing diethylhexylphthalate (DEHP), a compound transformed in vivo to metabolites known to activate PPARs, for 21 days (groups III and IV). Subsequently, animals were primed with Mycobacterium bovis purified protein derivative (PPD) in a Complete Freund's Adjuvant. Fifteen days later, animals in groups II and IV were challenged with Sepharose 4B beads covalently coupled with PPD, while animals in groups I and III received blank Sepharose beads. Animals with Th1 response (group II) showed a marked structural disruption in the hepatic lobule. Remarkably, these alterations were conspicuously absent in animals which received DEHP (group IV), despite noticeable accumulation of T cells in the periportal triads. Immunostaining and confocal microscopy revealed hepatic accumulation of IFNgamma+ Th1 and IL-4+ Th2 cells in animals from groups II and IV, respectively. Our data suggest a PPARalpha-mediated suppression of the development of a Th1 immune response in the liver, resulting in hepatoprotective effect. However, potentially negative consequences of PPAR activation, such as decreased ability of the immune system to fight infection and interference with the efficacy of vaccines designed to evoke Th1 immune responses, remain to be investigated.

Neutrophil-anti-microbial interaction in the established infection: effect on Staphylococcus aureus.

OBJECTIVES: To determine if differences in drug-related Staphylococcus aureus killing, associated in vivo with neutropenia, is neutrophil-related in vitro, and the mechanisms of this interaction. METHODS: To evaluate the influence of living neutrophils on drug-S. aureus interactions, cell wall enzymes, the PBPs, were isolated and their binding to five (beta lactam and other) antibiotics was evaluated following incubation (or not) with neutrophils. S. aureus killing by the test drugs was assayed in growth media of sterile filtered abscess fluid, either neutropenic infected or normal infected. At MBCs for the test isolate, each drug or saline control was incubated with S. aureus 10(6)and dilution-plated. RESULTS: Neutrophil incubation with S. aureus eliminated the S. aureus PBP-2 band in all Western blots irrespective of the drug used to tag the PBPs. Time-kill of S. aureus grown in neutropenic or normal abscess fluid showed greater kill by all drugs in neutropenic abscess fluid (p=0.029 6h incubation). Killing difference between the media correlates with drug PBP-2 activity. CONCLUSIONS: Drug activity against S. aureus in vitro is changed by neutrophil incubation. The neutrophil-induced loss of S. aureus PBP-2 drug binding suggests novel host-bacterial interaction that may impinge on drug treatment of S. aureus infections.

Transplant-related bronchiolitis obliterans (BOS) demonstrates unique cytokine profiles compared to toxicant-induced BOS.

Bronchiolitis obliterans (BOS - bronchiolitis obliterans syndrome - clinical diagnosis; CBO-histopathologic diagnosis), is a chronic disease process of fibrosis and cellular deposition in airways, complicating long term survival following lung transplantation. BOS is also the result of sporadic toxicant exposure, with airway signs, symptoms, and histology indistinguishable from allograft rejection. This study establishes a transplant BOS model in MHC-mismatched rats and compares their cytokine profiles and histopathology to that of our established toxicant-induced BOS model. Both models result in lung histopathology similar to human disease. Cytokines and inflammation markers that are elevated in human transplant BOS (TGFbeta, iNOS, IFNgamma) were also elevated significantly in both models. Anti-nuclear antibody was absent from all sera in transplant or toxicant models exhibiting advanced airway pathology. The cytokine osteopontin was highly elevated in BAL early in toxicant-induced BOS, but increased late in the transplant-induced BOS model. The data show that BOS is a disease of a pathologic endpoint that is induced by different triggers and processes. The highly elevated BAL osteopontin early in the toxicant-induced BOS model suggests a need for evaluation in the diagnostic setting.

The effect of treatment with a protease inhibitor on mycobacterial infection.

Mycobacterial infection occurs frequently in patients that receive protease inhibitors, which are drugs used to treat AIDS, but are known for metabolic effects. Proteases of microbial antigens have been recognized as important regulators of host inflammation and cellular response. To evaluate protease inhibitor effect on a mycobacterial infection, a pilot animal model was established. Mycobacterium bovis (bacillus Calmette-Guerin, or BCG) infection was compared in rats that received ritonavir and those that did not. Tissues and serum from one drug-treated and one control were analyzed weekly. Fewer acid-fast bacilli (AFBs) were consistently found in the drug-treated group by 3 separate measures: culture of tissue homogenates on solid media, tissue granuloma counts on organ sections, and staining of tissues for AFBs. Possible mechanisms of the observed relative resistance to BCG infection in ritonavir-treated rats were explored, by evaluating M. bovis cell wall lipids and proteins and by measuring infection-related cytokines in treated and control animals.

Helicobacter pylori-related immunoglobulins in sarcoidosis.

BACKGROUND/PURPOSE: The purpose of this study was to determine serum antibody titers against a common bacterial antigen, Helicobacter pylori (H. pylon), in subjects with sarcoidosis, comparing those titers to those present in a healthy population. SUBJECTS AND METHODS: With the approval of the Institutional Review Board of the University of Missouri-Kansas City, patients with sarcoidosis (pulmonary and extrapulmonary) who visited the Truman Medical Center-Hospital Hill pulmonary clinic were recruited to enter the study. A serum sample was frozen at -70 degrees C for later testing (n = 20). Specific information collected on subjects included corticosteroid use, use of histamine2 blockers and antacids, date of first diagnosis, and stage of sarcoidosis. Normal controls and demographically matched individuals who lacked pulmonary diseases, including sarcoidosis, were also recruited. Serum samples were processed as above. Antibody capture enzyme immunoassay was completed for H. pylori and urease antigens by serum dilution assay for each subject, from which titers for antigen-specific immunoglobulin (Ig)G and IgA were calculated. Nonspecific serum IgE was also measured. RESULTS: An increased incidence of high-titer IgG antibody directed against H. pylori antigens was found in subjects with sarcoidosis compared with controls. The sarcoidosis and control groups were significantly different with respect to IgG and IgA against H. pylori, both at p = .001. IgG directed against urease was also significantly different between sarcoidosis and control patients (p = .001), but IgA directed against urease was very low in all subjects and did not yield significant differences between groups. CONCLUSIONS: Specific H. pylori and urease IgG antibodies exceeded those expected in the population studied. The data suggest that in pulmonary sarcoidosis, the relationship of H. pylori and its products to sarcoid granuloma formation warrants further investigation.