Tetracyclines and pulmonary inflammation.

Tetracycline and its derivatives, such as chlortetracycline, oxytetracycline, minocycline, doxycycline, methacycline and lymecycline, are naturally occurring or semi-synthetic polyketide compounds that exhibit a well known broad-spectrum antibacterial activity that interferes with prokaryotic protein synthesis at the ribosome level. In addition to this well known antibacterial activity these compounds also exhibit a variety of additional, less well known properties. Among them are separate and distinct anti-inflammatory properties. Tetracycline and related compounds have been shown to be effective chemotherapeutic agents in a wide variety of chronic inflammatory diseases and conditions. These include periodontitis, rosacea, acne, auto-immune diseases such as rheumatoid arthritis and protection of the central nervous system against trauma and neurodegenerative diseases such as stroke, multiple sclerosis and Parkinson disease. Tetracycline and related compounds appear to be beneficial for treatment of several chronic inflammatory airway diseases. Among them are asthma, bronchiectasis, acute respiratory distress syndrome, chemical induced lung damage and cystic fibrosis. The clinical use of tetracycline-type drugs in treatment of chronic airway inflammation is becoming a topic of intense interest. Recent findings in this area have led to an understanding of the myriad physiological, cellular and molecular mechanisms of the inflammatory response and how this response may be controlled to limit damage to host cells and tissues. This review presents a brief summary of the recent research in the area of tetracycline and its derivatives in control of pulmonary inflammation.

Macrolide antibiotics and pulmonary inflammation.

Many clinically effective therapeutic agents can exhibit localized and systemic effects that are manifestly different from their intended primary pharmacological mode of action. Macrolide antibiotics such as erythromycin and its derivatives are no exception. In addition to their antibacterial action, this class of antibiotics exhibits anti-inflammatory activity in a variety of airway diseases such as asthma and diffuse panbronchiolitis that is separate and distinct from a direct antibacterial action. A variety of erythromycin derivatives have been shown to be clinically beneficial in these airway diseases. The anti-inflammatory activities of these macrolide antibiotics are becoming a research topic of intense interest. Recent work in this field has led to the understanding of the various physiological, cellular and molecular processes of the inflammatory response that are inhibited or suppressed by these compounds. This review presents a brief summary of the fascinating recent work in this active research area.

Temporal stability of the Minnesota Multiphasic Personality Inventory (MMPI) in patients undergoing lumbar fusion: a poor predictor of surgical outcome.

In this prospective study, the Minnesota Multiphasic Personality Inventory (MMPI) was administered to patients before and after lumbar spines fusion to investigate the stability of MMPI scores after surgical intervention and to attempt to correlate MMPI scale scores with outcome data. Sixty-eight patients were included. Testing was performed before surgery and at a mean of 1.5 years after surgery. Clinical outcome ratings were assigned by using criteria of pain relief and analgesic use. In addition, demographic variables known to affect outcome were analyzed. Sixty percent of the patients had a successful clinical outcome. Positive outcome correlated with the demographic factors of occupation (homemaker) and solid fusion. MMPI scales were stable across time, with no difference between groups. Independent t tests were used to study preoperative MMPI scores with respect to clinical outcome. Unsatisfactory outcomes were associated with higher scores on scales 1, 3, and 10 before surgery. Postoperative testing revealed significant outcome correlations--higher scores on scales 1, 2, 3, 5, 7, and 8 associated with an unsatisfactory outcome. However, discriminant function analysis of preoperative MMPI data was able to classify outcomes correctly in only 58.8% of the cases. The utility of the MMPI as a predictor of outcome after surgical intervention is quite limited. Use of group data and testing before and after surgery does not appear to influence this conclusion. Although the scores as a group were stable across time, the amount of variance in outcome that could not be accounted for by using MMPI scales as predictors was unacceptably large.

Peroxynitrite inhibits inducible (type 2) nitric oxide synthase in murine lung epithelial cells in vitro.

Peroxynitrite, formed by nitric oxide (NO) and superoxide, can alter protein function by nitrating amino acids such as tyrosine, cysteine, trytophan, or methionine. Inducible nitric oxide synthase (Type 2 NOS or iNOS) converts arginine to citrulline, releasing NO. We hypothesized that peroxynitrite could function as a negative feedback modulator of NO production by nitration of iNOS. Confluent cultures of the murine lung epithelial cell line, LA-4 were stimulated with cytokines to express iNOS, peroxynitrite was added, and the flasks sealed. After 3 h, NO in the headspace above the culture was sampled. Peroxynitrite caused a concentration-dependent decrease in NO. Similar results were obtained when 3-morpholinosydnonimine (SIN-1), a peroxynitrite generator, was added to the flasks. PAPA-NONOate, the NO generator, did not affect the headspace NO. Nitration of the iNOS was confirmed by detection of 3-nitrotyrosine by Western blotting. These data suggest a mechanism for inhibition of NO synthesis at inflammatory sites where iNOS, NO, and superoxide would be expected.

Methotrexate stimulates lung fibroblasts and epithelial cells to release eosinophil chemotactic activity.

OBJECTIVE: To determine if methotrexate (MTX) induces human lung fibroblast (HFL-1) and epithelial (BEAS 2B) cell lines to release eosinophil chemotactic activity (ECA). METHODS: HFL-1 and BEAS 2B cell supernatant fluids were evaluated for ECA by a blind well chamber technique. RESULTS: HFL-1 and BEAS-2B cells released ECA in a dose and time dependent manner in response to MTX. Partial characterization revealed that ECA was partly heat labile, trypsin sensitive, and ethylacetate extractable. Thus the culture supernatant fluids were evaluated for known eosinophil chemotactic factors. Although several were released constitutively, granulocyte-macrophage colony-stimulating factor (GM-CSF) was significantly increased in response to MTX from both cell types. Consistent with these observations, ECA from both cell types was inhibited by GM-CSF antibodies. CONCLUSION: These data suggest that lung fibroblasts and epithelial cells may modulate eosinophil recruitment into the lung by releasing ECA in response to MTX.

Inflammatory cytokines modulate eotaxin release by human lung fibroblast cell line.

Eotaxin, a potent eosinophil-specific chemotactic factor, is increased in the lower respiratory tract of asthma patients. Recently, lung fibroblasts have been reported to produce eotaxin and their activation can be modulated by inflammatory cytokines. To test the hypothesis that inflammatory cytokines modulate the eotaxin release from lung fibroblasts, we investigated the potential of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), or interferon-gamma (IFN-gamma) to induce the release of eotaxin and eotaxin mRNA by the human fetal lung fibroblast cell line, HFL-1, was evaluated. HFL-1 released eotaxin constitutively without stimulation, but IL-1beta or TNF-alpha stimulated eotaxin release in a dose- and time-dependent manner. IL-1beta or TNF-alpha treatment of HFL-1 also resulted in the augmented expression of eotaxin mRNA. Although IFN-gamma alone had negligible effect on eotaxin release and mRNA expression, IFN-gamma induced a significant, concentration-dependent attenuation of eotaxin release and eotaxin mRNA expression from HFL-1 stimulated with IL-1beta or TNF-alpha. These findings are consistent with the concept that lung fibroblast-derived eotaxin may in part be responsible for the eosinophil infiltration observed in the airways of asthmatic patients and that network of cytokines may modulate the eosinophil recruitment to the airways by stimulation of fibroblasts to release eotaxin.

Quality of life in ALS is maintained as physical function declines.

OBJECTIVES: To study patients with ALS to determine how physical function, quality of life (QOL), and spirituality or religiousness change over time, and what relationship these changes have to one another. METHODS: Sixty patients with ALS were studied prospectively. They were assessed at baseline, 3 months, and 6 months, using questionnaires designed to measure general quality of life (McGill Quality of Life questionnaire), religiosity (Idler Index of Religiosity), ALS-specific health-related quality of life (SIP/ALS-19), and ALS-specific function (ALS functional rating scale). RESULTS: A two-way repeated measures multivariate analysis of variance revealed that both the passage of time and the specific QOL scales used were factors in predicting patient quality of life (F[1, 59]= 9.87, p < 0.003 and F[3, 177]= 16.90, p < 0.001) Despite a progressive decline in physical function as measured by the ALS-specific function score, the general QOL and religiosity scores changed little. In contrast, the ALS-specific health-related QOL score declined in parallel with the ALS-specific function score. CONCLUSIONS: QOL in patients with ALS appears to be independent of physical function, which agrees with a previous cross-sectional study. The ALS-specific health-related QOL score is primarily a measure of physical function. QOL instruments that assess spiritual, religious, and psychological factors produce different results than those obtained using measures of physical function alone.

Erythromycin modulates eosinophil chemotactic cytokine production by human lung fibroblasts in vitro.

Recent studies suggest that erythromycin can suppress the production of some cytokines and may be an effective treatment for asthma. Eosinophil chemotactic cytokines have been suggested to contribute to the pathogenesis of asthma by the recruitment of eosinophils. We hypothesized that erythromycin modulates eosinophil chemotactic cytokine production. To test the hypothesis, we evaluated the potential of erythromycin to modulate the release of eosinophil chemoattractants from the human lung fibroblast cell line HFL-1. HFL-1 released eotaxin, granulocyte-macrophage colony-stimulating factor, and regulated and normal T-cell expressed and presumably secreted (RANTES) in response to interleukin-1beta or tumor necrosis factor alpha. Erythromycin attenuated the release of these cytokines and eosinophil chemotactic activity by the HFL-1. The suppressive effect on eotaxin was the most marked of these cytokines. Erythromycin therapy also suppressed eotaxin mRNA significantly. These results suggest a mechanism that may account for the apparent beneficial action of macrolide antibiotics in the treatment of allergic airway disorders.

Reactive oxygen and nitrogen metabolites modulate fibronectin-induced fibroblast migration in vitro.

Nitration of proteins by peroxynitrite may alter protein function. We hypothesized that reactive nitrogen species modulate fibronectin-induced fibroblast migration. To test this hypothesis, we evaluated fibroblast migration induced by fibronectin incubated with and without peroxynitrite. Peroxynitrite attenuated fibronectin-induced fibroblast migration in a dose-dependent manner but did not attenuate complement-activated serum-induced fibroblast migration. The reducing agents, deferoxamine and dithiothreitol (DTT), and L-tyrosine reversed the inhibition by peroxynitrite. PAPA-NONOate, a nitric oxide (NO) donor, and superoxide generated by the action of xanthine oxidase on lumazine or xanthine, also showed an inhibitory effect on fibroblast migration. The peroxynitrite generator, 3-morpholinosydnonimine (SIN-1), caused a concentration-dependent inhibition of fibroblast migration. Peroxynitrite reduced fibronectin binding to fibroblasts and resulted in nitrotyrosine formation. These findings are consistent with nitration of tyrosine by peroxynitrite with subsequent inhibition of fibronectin binding to fibroblasts and suggest that peroxynitrite may play a role in regulation of fibroblast migration.

Chicken soup inhibits neutrophil chemotaxis in vitro.

Chicken soup has long been regarded as a remedy for symptomatic upper respiratory tract infections. As it is likely that the clinical similarity of the diverse infectious processes that can result in "colds" is due to a shared inflammatory response, an effect of chicken soup in mitigating inflammation could account for its attested benefits. To evaluate this, a traditional chicken soup was tested for its ability to inhibit neutrophil migration using the standard Boyden blindwell chemotaxis chamber assay with zymosan-activated serum and fMet-Leu-Phe as chemoattractants. Chicken soup significantly inhibited neutrophil migration and did so in a concentration-dependent manner. The activity was present in a nonparticulate component of the chicken soup. All of the vegetables present in the soup and the chicken individually had inhibitory activity, although only the chicken lacked cytotoxic activity. Interestingly, the complete soup also lacked cytotoxic activity. Commercial soups varied greatly in their inhibitory activity. The present study, therefore, suggests that chicken soup may contain a number of substances with beneficial medicinal activity. A mild anti-inflammatory effect could be one mechanism by which the soup could result in the mitigation of symptomatic upper respiratory tract infections.

Quality of life in ALS depends on factors other than strength and physical function.

OBJECTIVES: To study patients with ALS to determine the following: 1) the relationship between physical function and quality of life (QOL); 2) the instruments that best reflect patients' own ratings of QOL; and 3) whether spiritual/religious factors play a role in determining QOL. METHODS: The authors prospectively studied 96 patients with ALS using several instruments, including the McGill Quality of Life (MQOL) instrument, the Idler Index of Religiosity, the Sickness Impact Profile (SIP)/ALS-19, and several measures of strength and physical function. RESULTS: QOL as assessed by patients (MQOL single item score) did not correlate with measures of physical function and strength, but correlated with the total MQOL score (p < 0.0005), the psychological and existential subscores of MQOL (p < 0. 0005), the support subscore of MQOL (p = 0.001), and the total Idler score (p = 0.001). In contrast, correlations between SIP/ALS-19 and these measures were not significant, although SIP/ALS-19 correlated with measures of physical function and strength. CONCLUSIONS: QOL, as assessed by the patient with ALS, does not correlate with measures of strength and physical function, but appears to depend on psychological and existential factors, and thus may be measured well by the MQOL scale. Spiritual factors and support systems appear to play roles as well. SIP/ALS-19 is a good measure of physical function, but not of overall QOL.

Elevation of nitrotyrosine and nitrate concentrations in cystic fibrosis sputum.

Nitric oxide (NO) is increased in the exhaled air of some patients with inflammatory lung disorders, but not in others. NO may combine with superoxide to form peroxynitrite, which lowers NO gas concentrations, increases formation of nitrate, and increases nitration of tyrosine residues on proteins. We hypothesized that superoxide released from neutrophils in the lower respiratory tract of cystic fibrosis (CF) results in increased nitrate and nitrotyrosine levels in sputum. In order to test this hypothesis, exhaled NO was collected from 5 stable adult CF subjects and from 5 nonsmoking normal controls. Consistent with previous reports, exhaled NO concentrations were not increased in CF exhaled air (22.6 +/- 1.5 ppb vs. 28.6 +/- 1.5 ppb in normals, P > 0.05). Sputum was collected from 9 adult CF subjects and the same 5 normal controls and evaluated for nitrite, nitrate, and nitrotyrosine. Nitrate and nitrotyrosine levels, but not nitrite, were significantly elevated in CF. Recently, myeloperoxidase has also been implicated as a mechanism of nitrotyrosine formation. Therefore, myeloperoxidase was measured and found to be elevated in the CF sputum (64.2 +/- 35.9 vs. 0.73 +/- 0.16 U/mL, P < 0.001), and was found to correlate with concentrations of nitrotyrosine (r = 0.87, P < 0.05). However, in vitro studies with myeloperoxidase and murine lung epithelial cells did not demonstrate a reduction of NO gas with nitrotyrosine or an increase in nitrate formation. These data demonstrate that nitrate and nitrotyrosine are elevated in the sputa of CF subjects and suggest increased production of NO in the lower respiratory tract of CF patients, despite the relatively low exhaled NO levels. Pediatr Pulmonol. 2000; 30:79-85. Published 2000 Wiley-Liss, Inc.

Bradykinin stimulates eotaxin production by a human lung fibroblast cell line.

BACKGROUND: Bradykinin, a potent inflammatory peptide, is increased in the airways of allergic patients. Accompanying the elevated bradykinin levels are increases in both eosinophils and fibroblasts. Eotaxin, a potent eosinophil-specific chemotactic factor, is released by fibroblasts and increased in the lower respiratory tract of allergic patients. OBJECTIVE: We sought to test the hypothesis that lung fibro-blasts release eotaxin in response to bradykinin. METHODS: The potential of bradykinin to induce the release of eotaxin from the human lung fibroblast cell line HFL-1 was tested by cell culture and evaluation of the culture supernatant fluids and RNA for immunoreactive eotaxin and eotaxin messenger RNA. RESULTS: HFL-1 cells released eotaxin constitutively without stimulation, but bradykinin stimulated eotaxin release in a dose- and time-dependent manner and resulted in augmented expression of eotaxin messenger RNA. The release of eotaxin was sensitive to the action of glucocorticoids. Eosinophil chemotactic activity by HFL-1 supernatant fluids was inhibited by anti-human eotaxin-neutralizing antibody. Consistent with these results, inhibitors of bradykinin B2 receptors, but not bradykinin B1 receptors, inhibited bradykinin-induced eotaxin release. CONCLUSION: These data demonstrate that bradykinin may stimulate lung fibroblasts to release eotaxin and suggest the potential for this mechanism to be important in modulation of lung inflammation.

Endogenous nitric oxide in allergic airway disease.

There has been intense research into the role nitric oxide (NO) plays in physiologic and pathologic mechanisms. The presence of NO in exhaled breath and the high concentrations in nasal airways stimulated many studies examining exhaled and nasal NO as potential markers of airway inflammation, enabling repeated monitoring of airway inflammation not possible with invasive tests (eg, bronchoscopy). In airway inflammation, NO is not merely a marker but may have anti-inflammatory and proinflammatory effects. Nasal NO measurement may be used in the noninvasive diagnosis and monitoring of nasal disease. This review was compiled by speakers who gave presentations on NO at the annual meeting of the American Academy of Allergy, Asthma, and Immunology in 1999 on exhaled and nasal NO, in vitro studies of NO, the chemistry of airway NO formation, and standardized measurement of exhaled mediators.

Inhibition of MIP-1alpha-induced human neutrophil and monocyte chemotactic activity by reactive oxygen and nitrogen metabolites.

Peroxynitrite, formed by the reaction between nitric oxide (NO) and superoxide, has been implicated in the pathogenesis of numerous disease processes. Several studies have shown that peroxynitrite-induced protein nitration may compromise enzyme and protein function. We hypothesized that peroxynitrite may regulate cytokine function during inflammation. To test this hypothesis, the neutrophil and monocyte chemotactic responses of macrophage inflammatory protein-1alpha (MIP-1alpha) incubated with and without peroxynitrite were evaluated. Peroxynitrite attenuated neutrophil chemotactic activity (NCA) and monocyte chemotactic activity (MCA) by MIP-1alpha in a dose-dependent manner (P < .05). The inhibitory effects were not significant on NCA and MCA induced by leukotriene B4 or complement-activated serum incubated with peroxynitrite. The reducing agents deferoxamine, dithiothreitol, and exogenous L-tyrosine abrogated the NCA and MCA inhibition by peroxynitrite. Papa-NONOate, an NO donor, or a combination of xanthine and xanthine oxidase to generate superoxide, did not show an inhibitory effect on NCA and MCA induced by MIP-1alpha. In contrast, 3-morpholinosydnonimine (SIN-1), a peroxynitrite generator, elicited a concentration-dependent reduction in NCA and MCA induced by MIP-1alpha. Consistent with its capacity to reduce NCA and MCA, peroxynitrite treatment reduced MIP-1alpha binding to neutrophils and monocytes. Nitrotyrosine was detected in the MIP-1alpha incubated with peroxynitrite. These findings are consistent with nitration of tyrosine by peroxynitrite with subsequent inhibition of MIP-1alpha binding to neutrophils and monocytes and a reduction in NCA and MCA. These data demonstrate that peroxynitrite modulates the inflammatory cell migration by MIP-1alpha, and they suggest that oxidants may play an important role in the regulation of MIP-1alpha-induced inflammatory cell chemotaxis.

Bleomycin stimulates lung fibroblast and epithelial cell lines to release eosinophil chemotactic activity.

The presence of eosinophils in the lungs of patients with pulmonary fibrosis correlates with poor prognosis or resistance to therapy. Furthermore, eosinophils localize to areas undergoing active fibrosis. It was hypothesized that a human lung fibroblast (HFL-1) and a human lung epithelial cell line (BEAS-2B) might release eosinophil chemotactic activity (ECA) in response to bleomycin, a chemotherapeutic agent associated with pulmonary fibrosis. HFL-1 and BEAS-2B cells were cultured in the presence of bleomycin and their supernatant fluids evaluated for ECA by means of a Boyden chamber method. HFL-1 and BEAS-2B cells released ECA in a dose- and time-dependent manner in response to bleomycin, and partial characterization revealed that the ECA was heterogeneous. ECA release from HFL-1 and BEAS-2B cells was significantly reduced by a leukotriene B4 (LTB4) receptor antagonist and an antibody directed against granulocyte-macrophage colony-stimulating factor. HFL-1 cells released LTB4, eotaxin, and GM-CSF constitutively, and BEAS-2B cells released LTB4, eotaxin, regulated on activation, normal T-cell expressed and presumably secreted, and GM-CSF constitutively. In both cases, the release of GM-CSF was significantly increased in response to bleomycin. These data suggest that lung fibroblasts and epithelial cells may modulate eosinophil recruitment into the lung in bleomycin-induced pulmonary fibrosis.

Reactive nitrogen and oxygen species attenuate interleukin- 8-induced neutrophil chemotactic activity in vitro.

Peroxynitrite, formed by the reaction between nitric oxide and superoxide, has been shown to induce protein nitration, which compromises protein function. We hypothesized that peroxynitrite may regulate cytokine function during inflammation. To test this hypothesis, the neutrophil chemotactic activity (NCA) of interleukin-8 (IL-8) incubated with peroxynitrite was evaluated. Peroxynitrite attenuated IL-8 NCA in a dose-dependent manner (p < 0.01) but did not significantly reduce NCA induced by leukotriene B(4) or complement-activated serum. The reducing agents, dithionite, deferoxamine, and dithiothreitol, reversed and exogenous L-tyrosine abrogated the peroxynitrite-induced NCA inhibition. Papa-NONOate [N-(3-ammoniopropyl)-N-(n-propyl)amino]diazen-1-ium-1, 2-dialase or sodium nitroprusside, NO donors, or a combination of xanthine and xanthine oxidase to generate superoxide did not show an inhibitory effect on NCA induced by IL-8. In contrast, small amounts of SIN-1, a peroxynitrite generator, caused a concentration-dependent inhibition of NCA by IL-8. Consistent with its capacity to reduce NCA, peroxynitrite treatment reduced IL-8 binding to neutrophils. Nitrotyrosine was detected in the IL-8 incubated with peroxynitrite by enzyme-linked immunosorbent assay. These findings are consistent with nitration of tyrosine by peroxynitrite with subsequent inhibition of IL-8 binding to neutrophils and a reduction in NCA and suggest that oxidants may play an important role in regulation of IL-8-induced neutrophil chemotaxis.

Effects of reactive oxygen and nitrogen metabolites on eotaxin-induced eosinophil chemotactic activity in vitro.

Peroxynitrite, an oxidant generated by the interaction between superoxide and nitric oxide (NO), has been implicated in the etiology of numerous disease processes. Several studies have shown that peroxynitrite-induced protein nitration may compromise enzyme and protein function. We hypothesized that peroxynitrite may regulate cytokine function during inflammation. To test this hypothesis, the eosinophil chemotactic responses of eotaxin incubated with and without peroxynitrite were evaluated. Peroxynitrite attenuated eotaxin-induced eosinophil chemotactic activity (ECA) in a dose-dependent manner (P < 0.05). The inhibitory effects were not significant on ECA induced by leukotriene B(4) or complement-activated serum incubated with peroxynitrite. The reducing agents deferoxamine and dithiothreitol reversed the ECA inhibition by peroxynitrite, and exogenous L-tyrosine abrogated the inhibition by peroxynitrite. PAPA-NONOate (an NO donor) or a combination of xanthine and xanthine oxidase to generate superoxide did not show an inhibitory effect on ECA induced by eotaxin. In contrast, 3-morpholinosydnonimine, a peroxynitrite generator, caused a concentration-dependent inhibition of ECA by eotaxin. Consistent with its capacity to reduce ECA, peroxynitrite treatment reduced eotaxin binding to eosinophils. Nitrotyrosine was detected in the eotaxin incubated with peroxynitrite. These findings are consistent with nitration of tyrosine by peroxynitrite with subsequent inhibition of eotaxin binding to eosinophils and a reduction in ECA. These data demonstrate that peroxynitrite modulates the eosinophil migration by eotaxin, and suggest that oxidants may play an important role in regulation of eotaxin-induced eosinophil chemotaxis.