![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
Previous Article | Next Article 
Infection and Immunity, July 2002, p. 3649-3655, Vol. 70, No. 7
0019-9567/02/$04.00+0 DOI: 10.1128/IAI.70.7.3649-3655.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.
Division of Bacterial and Mycotic Diseases,1 Division of AIDS, Sexually Transmitted Diseases, and Tuberculosis Laboratory Research, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 303332
Received 25 January 2002/ Returned for modification 18 March 2002/ Accepted 18 April 2002
 |
ABSTRACT |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
INTRODUCTION |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Cytadherence of M. pneumoniae to the respiratory epithelium is regarded as an essential primary step in tissue colonization and subsequent disease pathogenesis (reviewed in reference 42 and references therein). The organism has developed a special organelle at the tip of the elongated flask-shaped cell that mediates attachment. Two surface proteins, the 170-kDa P1 and the 30-kDa P30 proteins, function as adhesins, while several other accessory proteins (HMW1, HMW2, and HMW3 and A, B, and C) collectively maintain the proper distribution and/or disposition of the adhesions in the mycoplasma membrane (3, 10, 27, 34). The effects of M. pneumoniae infection on the immune system also contribute to pathogenesis. High percentages of neutrophils and lymphocytes are present in the bronchoalveolar lavage fluid in patients with Mycoplasma pneumonia, and levels of IL-1ß, IL-2, IL-4, and IL-6 are elevated in both the bronchoalveolar lavage fluid and serum of these patients (26, 30).
Cytokines are important mediators in both lung defense and inflammation (24). Chen et al. (7-9) have shown that IL-1, IL-6, and TNF-
were induced in Pneumocystis carinii infection with a murine model, and these cytokines played important roles in host resistance to P. carinii by regulating the pulmonary inflammation responses, such as recruiting inflammatory cells into the lungs. On the other hand, Ulich et al. (48) reported that IL-6 and transforming growth factor ß could down-regulate and curtail the exodus of neutrophils into local acute inflammatory sites, thus suggesting the presence of an endogenous negative feedback mechanism to inhibit endotoxin-initiated cytokine-mediated acute inflammation. In addition to the observation of elevated cytokine levels in Mycoplasma pneumonia patients, M. pneumoniae was also shown to induce proinflammatory cytokine gene expressions in mouse models, including expression of IL-1ß, IL-6, TNF-, or IFN-
(37, 38). M. pneumoniae also has similar effects on human peripheral blood mononuclear cells or Epstein-Barr virus-positive lymphoblastoid cell lines (25, 44). These data all point to the possible role of proinflammatory cytokines in the pathogenesis of M. pneumoniae. Although respiratory epithelial cells are the main targets of M. pneumoniae, there are no reports of their involvement in the cytokine response to infection with this respiratory pathogen. Because epithelial cells at mucosal surfaces have been shown to secrete chemoattractant and proinflammatory cytokines in response to bacterial infections (13), we decided to investigate the regulation and mechanisms of the cytokine response in respiratory epithelial cells during M. pneumoniae infection.
|
MATERIALS AND METHODS |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Cell line and reagents. A549 cells (human lung epithelial carcinoma cells; American Type Culture Collection) were grown in Earle's Minimum Essential Medium (EMEM) with 10% fetal bovine serum (GIBCO, Grand Island, N.Y.). Cells were free of mycoplasmas as routinely determined by a Mycoplasma PCR Primer set (Stratagene, La Jolla, Calif.). For M. pneumoniae treatment, 500 µl of M. pneumoniae from the same frozen stock was always added to 5 ml of cell culture medium (1:10, vol/vol; appropriately 5 x 107 CFU/106 cells) unless otherwise specified. Cycloheximide (CH), chloramphenicol (CP), and nalidixic acid (NA) were all purchased from Sigma (St. Louis, Mo.). CH was prepared as a 1-mg/ml stock in EMEM and added to a final concentration of 10 µg/ml. CP was dissolved in absolute ethanol as a 10-mg/ml stock and added to a final concentration of 20 µg/ml. NA was dissolved in EMEM as a 5-mg/ml stock and added to a final concentration of 40 µg/ml.
Cell viability assay. The CellTiter 96 Aqueous One Solution Cell Proliferation assay (Promega, Madison, Wis.) is a colorimetric method for determining the number of viable cells in proliferation or cytotoxicity assays. It was performed as previously described (52). Briefly, 104 cells were seeded into a 96-well culture plate (Corning, Corning, N.Y.) and treated with M. pneumoniae for specified times. At the end of treatment, 20 µl of the CellTiter solution was added directly to each culture well and incubated at 37°C for 2 h, when the absorbance at 490 nm was determined with a Bio-Rad (Hercules, Calif.) microplate reader.
For cell counting, 105 cells were seeded into 12-well plates (Corning). After treatment, cells were harvested with 0.25% trypsin, and the number of trypan blue-excluding cells was determined with a hemacytometer.
Cytokine assays.
A total of 5 x 105 A549 cells were seeded into a six-well plate and treated with M. pneumoniae for up to 24 h. Cytokines secreted into the culture supernatant were detected using enzyme-linked immunosorbant assays (ELISAs). An OptEIA Human IL-1ß set was used to detect IL-1ß (15.6 to 1,000 pg/ml) (BD Biosciences, San Diego, Calif.), and the DuoSet ELISA Development systems were used to detect IL-6 (4.2 to 300pg/ml), IL-8 (31.25 to 2,000 pg/ml), IFN- (15.6 to 1,000 pg/ml), and TNF- (15.6 to 1,000 pg/ml) (R & D Systems, Minneapolis, Minn.). All assays were conducted according to the manufacturers' instructions.
Immunofluorescent microscopy. To detect intracellular IL-1ß in A549 cells, an indirect immunofluorescent method was used. A total of 104 cells were seeded into a 96-well plate. At 24 h postinfection (PI), the culture medium was removed, and 200 µl of ice-cold methanol was added to each well to fix and permeate the cells for 5 min at room temperature. After the methanol was removed, 25 µl of mouse anti-human IL-1ß antibody (dilution of 1:250; BD Biosciences) was added to each well and incubated at 37°C for 30 min in the dark. The antibodies were removed, and wells were washed twice with phosphate-buffered saline (0.01 M; pH 7.2). Twenty-five microliters of fluorescein isothiocyanate (FITC)-labeled goat anti-mouse antibody (1:1,000; BD Biosciences) was then added to each well and incubated at 37°C for another 30 min in the dark. The secondary antibody was removed, the wells were washed with phosphate-buffered saline, and then mounting fluid (carbonate-buffered glycerol mounting fluid; pH 9.0) was added. The plate was evaluated with an Olympus IX70 fluorescent microscope. Images were captured and analyzed with the Optronics (Goleta, Calif.) program.
RNA preparation and quantification. Total RNA was prepared from A549 cells with the RNeasy Mini kit (Qiagen, Valencia, Calif.). Briefly, medium was completely removed from the culture flasks, and then 350 µl of RLT lysis buffer (Qiagen) was added into each 25-cm2 flask to lyse the cells. After the lysate was transferred to a 1.5-ml collection tube, 350 µl of 70% ethanol was added and mixed well by repeated pipetting. The mixture was again transferred to an RNeasy mini spin column and centrifuged for 15 s at 8,000 x g. The column was then washed with RW1 and RPE buffer (Qiagen), and finally the RNA was dissolved in 30 µl of DNase- and RNase-free water. The quantity and purity of the RNA samples were determined by measuring the absorbance at 260 nm and the 260 nm/280 nm absorbance ratio with a spectrometer.
RT-PCR.
Ready-To-Go RT-PCR beads (Amersham Pharmacia Biotech Inc., Piscataway, NJ) were used for the reverse transcription (RT)-PCR assay. Total RNA (0.5 µg) was added to each reaction mixture. The primer sequences were as follows: IL-1ß, 5'-AAA CAG ATG AAG TGC TCC TTC CAG G-3' and 5'-TGG AGA ACA CCA CTT GTT GCT CCA-3' (16); IL-6, 5'-GGC TGA AAA AGA TGG ATG CT-3' and 5'-CCT GCT TCA CCA CCT TCT G-3' (53); IL-8, 5'-AGA TAT TGC ACG GGA GAA-3' and 5'-GAA ATA AAG GAG AAA CCA-3'; ß-actin, 5'-CGG GAC CTG ACT GAC TAC-3' and 5'-GAA GGA AGG CTG GAA GAG-3' (31); IFN-, 5'-GAT GCT CTT CGA CCT TGA AAC AGC AT-3' and 5'-ATG AAA TAT ACA AGT TAT ATC TTG GCT TTT-3' (15); and TNF-, 5'-GAG TGA CAA GCC TGT AGC CCA TGT TGT AGC A-3' and 5'-GCA ATG ATC CCA AAG TAG ACC TGC CCA GAC T-3' (49). The RT was carried out at 42°C for 30 min, and the amplifications were carried out for 32 cycles of 30 s at 95°C, 30 s at 55°C, and 60 s at 72°C. The products were run on a 2% agarose gel and stained with ethidium bromide. The gels were then scanned with a FluorImager SI (Molecular Dynamics, Sunnyvale, Calif.) and quantified using the ImageQuant program, version 4.2A (Molecular Dynamics), and the percentage of surface area under the peak of each band was normalized to that of the corresponding ß-actin band.
Adherence blockage assay. A mouse anti-M. pneumoniae monoclonal antibody (immunoglobulin G2a [IgG2a] isotype; Chemicon International, Inc., Temecula, Calif.) which targets the P1 adhesin protein was incubated with M. pneumoniae stock solution at a 1:10, 1:100, or 1:1,000 ratio for 2 h at room temperature. The cells were then inoculated with the mixture, and after 4 h the total RNA was extracted and assayed for IL-1ß mRNA. A purified mouse IgG2a antibody with an unrelated epitope was used as an isotype control (Caltag).
Protease digestion treatment. Protease digestion was conducted as described by Hu et al. (22) with some modifications. In some experiments, M. pneumoniae was pretreated with trypsin (GIBCO) for 20 min at 37°C at a final concentration of 100 µg/ml to remove surface adhesion proteins and was then used to infect A549 cells.
Statistical analysis. Statistical analysis was performed with Student's t test. A probability level of P < 0.05 was considered significant. Data are presented as the mean ± the standard deviation.
|
RESULTS |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
When tested by RT-PCR with ß-actin as an internal control, it was found that mycoplasma infection of A549 cells resulted in elevation of IL-1ß, IL-8, and TNF- mRNA (Fig. 1), whereas IL-6 and IFN- mRNA levels remained the same or undetectable (data not shown). IL-1ß mRNA was induced as soon as 1 h PI, with the signal becoming gradually stronger until 24 h PI. Occasionally, a faint nonspecific band below the IL-1ß band could be seen in IL-1 ß RT-PCR gels in the samples, but it did not interfere with the IL-1ß band. IL-8 had a rather strong baseline mRNA expression compared to IL-1ß, and the induction became obvious around 8 h PI. The induction of TNF- was the most transient; its mRNA appeared around 1 h PI, peaked around 4 to 8 h, and then rapidly decreased. There was a baseline expression of IL-6 mRNA, but it remained unchanged during infection (data not shown). IFN- mRNA, however, was not detected in either the control or the infected cells. The A549 cells cultured with SP4 medium had the same mRNA levels as the control cells in all experiments (data not shown). The three groups (A549 infected, A549 SP4 medium control, and untreated A549 cells) had the same levels of ß-actin mRNA expression during the 24-h period (Fig. 1 and data not shown).
|
have different protein expression patterns.
Because IL-1ß, IL-8, and TNF- mRNA levels were increased after M. pneumoniae infection, the levels of protein expression for these cytokines were investigated. Cell culture supernatants were collected at specific time points and analyzed for cytokine expression by ELISAs. As shown in Fig. 2A, it was found that IL-8 protein levels steadily increased during the 24-h infection period. TNF- protein was also expressed, but unlike IL-8, expression reached its peak just hours after infection and then started decreasing around 12 h PI (Fig. 2B). ELISA detected no IL-6 expression, as expected (data not shown).
|
|
|
A549 cell protein synthesis is required for increased cytokine levels. To examine whether the increase in cytokine levels was due to de novo protein synthesis, CH was added to the cell culture for 2 h before M. pneumoniae was added. At specific time points, culture medium was removed and IL-8 levels were measured. Although CH treatment did not affect the increased expression of cytokine genes (using IL-1ß as an indicator) (Fig. 5A), it did inhibit the induction of IL-8 protein compared to that with M. pneumoniae infection alone (Fig. 5B). These data strongly suggested that de novo synthesis is required for the increased levels of proinflammatory cytokines.
|
|
DISCUSSION |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Although we detected the induction of IL-8 and TNF- proteins in the cell culture medium by ELISA (Fig. 2), we did not detect the presence of any IL-1ß protein in the culture medium. IL-1, unlike other cytokines, lacks the typical hydrophobic signal peptide (5, 23, 50). The mature form of IL-1ß is a 17-kDa protein, which is processed from its 31-kDa (269-amino-acid) precursor (1). Intracellular IL-1ß consists exclusively of the 31-kDa precursor form, while extracellular IL-1ß consists of a mixture of both the unprocessed and mature forms of IL-1ß (19). Further studies revealed that even though several cell types have the capacity to produce the IL-1ß precursor, its release is predominantly limited to monocytes and macrophages (11). Therefore, by using an indirect immunofluorescent method, we did detect the presence of IL-1ß protein in M. pneumoniae-infected cells but not in control or mock-treated cells (Fig. 3). This provided clear evidence that IL-1ß protein was synthesized in human lung epithelial cells during the infection and that the protein remained intracellular instead of secreted. It is in consistent with a study conducted by Uchida et al.(47) in which it was shown that although IL-1ß mRNA in human gingival epithelial cells was induced in response to Actinobacillus actinomycetemcomitans, secretion of IL-1ß could not be detected. This is in direct contrast to some other reports that IL-1ß is secreted during infection (25, 44). Therefore, the ability of epithelial cells to secret IL-1ß protein may be very limited.
The mechanism for this limitation on the secretion may rely on the enzyme responsible for the processing of IL-1ß. The IL-1ß-converting enzyme (ICE; also called caspase 1) is able to process IL-1ß and is also involved in apoptosis (33, 36, 46). Hogquist et al. (20, 21) had shown that IL-1ß release is correlated with cell injury and that the processing of the precursor protein only occurs efficiently in cells that are undergoing apoptosis. However, we did not observe any significant cell damage in A549 cells during infection, and it is possible that the IL-1ß-converting enzyme was not activated and, thus, that no mature IL-1ß protein was secreted.
Several reports have demonstrated the induction of IL-6 in human lymphocytes, human peripheral blood mononuclear cells, and/or mice during M. pneumoniae infection (25, 37, 44). However, we did not detect the induction of IL-6 in the A549 human epithelial cell line following M. pneumoniae infection at the level of sensitivity of our ELISA. On the other hand, we did detect the induction of IL-6 in TNF--treated A549 cells (data not shown). However, Uchida et al. (47) had reported that IL-6 was not induced in human gingival epithelial cells in response to A. actinomycetemcomitans, but human gingival fibroblasts responded to A. actinomycetemcomitans by increasing their IL-6 mRNA levels. Therefore, it appears that various cell types in different models respond to the same infectious agent with different cytokine induction profiles.
Even though cytadherence appears to be important for cytokine induction, some other mechanisms are most likely to be involved. For example, it has been reported that bacterial protein synthesis is required for the induction of IL-8 by epithelial cells following chlamydial infection (39). Therefore, the possible roles of M. pneumoniae protein synthesis and DNA synthesis in cytokine induction were also evaluated. Our results (Fig. 5A) showed that these two macromolecule syntheses do not appear to be prerequisites for M. pneumoniae-induced cytokine gene expressions. However, A549 cellular protein synthesis is responsible for increased cytokine protein levels, as CH, an inhibitor of eukaryotic protein synthesis, can abolish the increase of IL-8 levels during M. pneumoniae infection (Fig. 5B).
This study presents a new perspective in M. pneumoniae pathogenesis, as it sheds light on the mechanisms of acute inflammation during infection. The association of this bacterium with chronic diseases, such as asthma and arthritis, and the mechanisms for host cell damage can be partly attributed to the induction of these proinflammatory cytokines during infection. It should be kept in mind that results from in vitro studies do not always mimic in vivo responses in the host. The serum in the cell culture medium may also influence the cytokine response to M. pneumoniae infection. The A549 cells we used are derived from a human lung epithelial carcinoma, and a cell line not associated with disease, such as primary lung epithelial cells, will be useful in future studies.
|
ACKNOWLEDGMENTS |
|---|
J. Yang is an American Society for Microbiology/National Center for Infectious Diseases Postdoctoral Research Associate.
|
FOOTNOTES |
|---|
|
REFERENCES |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
| 1. | Auron, P. E., S. J. Warner, A. C. Webb, J. G. Cannon, H. A. Bernheim, K. J. McAdam, L. J. Rosenwasser, G. LoPreste, S. F. Mucci, and C. A. Dinarello. 1987. Studies on the molecular nature of human interleukin 1. J. Immunol. 138:1447-1456.[Abstract] |
| 2. | Baseman, J. B., M. Lange, N. L. Criscimagna, J. A. Giron, and C. A. Thomas. 1995. Interplay between mycoplasma and host target cells. Microb. Pathog. 19:105-116.[CrossRef][Medline] |
| 3. | Baseman, J. B., and J. G. Tully. 1997. Mycoplasmas: sophisticated, reemerging, and burdened by their notoriety. Emerg. Infect. Dis. 3:21-32.[Medline] |
| 4. | Bernatchez, C., R. Al-Daccak, P. E. Mayer, K. Mehindate, L. Rink, S. Mecheri, and W. Mourad. 1997. Functional analysis of Mycoplasma arthritidis-derived mitogen interactions with class II molecules. Infect. Immun. 65:2000-2005.[Abstract] |
| 5. | Bjorkdahl, O., A. G. Wingren, G. Hedhund, L. Ohlsson, and M. Dohlsten. 1997. Gene transfer of a hybrid interleukin-1 beta gene to B16 mouse melanoma recruits leukocyte subsets and reduces tumour growth in vivo. Cancer Immunol. Immunother. 44:273-281.[CrossRef][Medline] |
| 6. | Bohnet, S., U. Kotschau, J. Braun, and K. Dalhoff. 1997. Role of interleukin-8 in community-acquired pneumonia: relation to microbial load and pulmonary function. Infection 25:95-100.[CrossRef][Medline] |
| 7. | Chen, W., E. A. Havell, F. Gigliotti, and A. G. Harmsen. 1993. Interleukin-6 production in a murine model of Pneumocystis carinii pneumonia: relation to resistance and inflammatory response. Infect. Immun. 61:97-102. |
| 8. | Chen, W., E. A. Havell, and A. G. Harmsen. 1992. Importance of endogenous tumor necrosis factor alpha and gamma interferon in host resistance against Pneumocystis carinii infection. Infect. Immun. 60:1279-1284. |
| 9. | Chen, W., E. A. Havell, L. L. Moldawer, K. E. McIntyre, R. A. Chizzonite, and A. G. Harmsen. 1992. Interleukin-1: an important mediator of host resistance against Pneumocystis carinii. J. Exp. Med. 176:713-718. |
| 10. | Dallo, S. F., A. L. Lazzell, A. Chavoya, S. P. Reddy, and J. B. Baseman. 1996. Bifunctional domains of the Mycoplasma pneumoniae P30 adhesin. Infect. Immun. 64:2595-2601.[Abstract] |
| 11. | Dinarello, C. A. 1991. Interleukin-1 and interkeukin-1 antagonism. Blood 77:1627-1652. |
| 12. | D'Orazio, J. A., B. C. Cole, and J. Stein-Streilein. 1996. Mycoplasma arthritidis mitogen up-regulates human NK cell activity. Infect. Immun. 64:441-447.[Abstract] |
| 13. | Eckman, L., M. F. Kagnoff, and J. Fierer. 1995. Intestinal epithelial cells as watchdogs for the natural immune system. Trends Microbiol. 3:118-120.[CrossRef][Medline] |
| 14. | Faulkner, C. B., J. W. Simecka, M. K. Davidson, J. K. Davis, T. R. Schoeb, J. R. Lindsey, and M. P. Everson. 1995. Gene expression and production of tumor necrosis factor alpha, interleukin 1, interleukin 6, and gamma interferon in C3H/HeN and C57BL/6N mice in acute Mycoplasma pulmonis disease. Infect. Immun. 63:4084-4090.[Abstract] |
| 15. | Fenton, M. J., M. W. Vermeulen, S. Kim, M. Burdick, R. M. Strieter, and H. Kornfeld. 1997. Induction of gamma interferon production in human alveolar macrophages by Mycobacterium tuberculosis. Infect. Immun. 65:5149-5156.[Abstract] |
| 16. | Gan, X., and B. Bonavida. 1999. Preferential induction of TNF- and IL-1ß and inhibition of IL-10 secretion by human peripheral blood monocytes by synthetic aza-alkyl lysophopholipids. Cell. Immunol. 193:125-133.[CrossRef][Medline] |
| 17. | Garcia, J., B. Lemercier, S. Roman-Roman, and G. Rawadi. 1998. A Mycoplasma fermentans-derived synthetic lipopeptide induces AP-1 and NF- B activity and cytokine secretion in macrophages via the activation of mitogen-activated protein kinase pathways. J. Biol. Chem. 273:34391-34398. |
| 18. | Hall, R. E., S. Agarwal, and D. P. Kestler. 2000. Induction of leukemia cell differentiation and apoptosis by recombinant P48, a modulin derived from Mycoplasma fermentans. Biochem. Biophys. Res. Commun. 269:284-289.[CrossRef][Medline] |
| 19. | Hazuda, D. J., J. C. Lee, and P. R. Young. 1988. The kinetics of interleukin 1 secretion from activated monocytes. Differences between interleukin 1 alpha and interleukin 1 beta. J. Biol. Chem. 263:8473-8479. |
| 20. | Hogquist, K. A., M. A. Nett, E. R. Unanue, and D. D. Chaplin. 1991. Interleukin 1 is processed and released during apoptosis. Proc. Natl. Acad. Sci. USA 88:8485-8489. |
| 21. | Hogquist, K. A., E. R. Unanue, and D. D. Chaplin. 1991. Release of IL-1 from mononuclear phagocytes. J. Immunol. 147:2181-2186.[Abstract] |
| 22. | Hu, P. C., A. M. Collier, and J. B. Baseman. 1977. Surface parasitism by Mycoplasma pneumoniae of respiratory epithelium. J. Exp. Med. 145:1328-1343. |
| 23. | Jarry, A., G. Vallette, E. Cassagnau, A. Moreau, C. Bou-Hanna, P. Lemarre, E. Letessier, J.-C. Le Neel, J.-P. Galmiche, and C. L. Laboisse. 1999. Interleukin1 and interleukin 1beta converting enzyme (caspase 1) expression in the human colonic epithelial barrier. Caspase 1 downregulation in colon cancer. Gut 45:246-251. |
| 24. | Kelly, J. 1990. Cytokines of the lung. Am. Rev. Respir. Dis. 141:765-788.[Medline] |
| 25. | Kita, M., Y. Ohmoto, Y. Hirai, N. Yamaguchi, and J. Imanishi. 1992. Induction of cytokines in human peripheral blood mononuclear cells by mycoplasmas. Microbiol. Immunol. 36:507-516.[Medline] |
| 26. | Koh, Y. Y., Y. Park, H. J. Lee, and C. K. Kim. 2001. Levels of interleukin-2, interferon-, and interleukin-4 in bronchoalveolar lavage fluid from patients with Mycoplasma pneumonia: implication of tendency toward increased immunoglobulin E production. Pediatrics 107:e39. |
| 27. | Krause, D. C. 1998. Mycoplasma pneumoniae cytadherence: organization and assembly of the attachment organelle. Trends Microbiol. 6:15-18.[CrossRef][Medline] |
| 28. | Krause, D. C., and J. B. Baseman. 1983. Inhibition of Mycoplasma pneumoniae hemadsorption and adherence to respiratory epithelium by antibodies to a membrane protein. Infect. Immun. 39:1180-1186. |
| 29. | Kunkel, S. L., T. J. Standiford, K. Kasahara, and R. M. Strieter. 1991. Interleukin-8 (IL-8): the major neutrophil chemotactic factor in the lung. Exp. Lung Res. 17:17-23.[Medline] |
| 30. | Lieberman, D., S. Livnat, F. Schlaeffer, A. Porath, S. Horowitz, and R. Levy. 1997. IL-1ß and IL-6 in community-acquired pneumonia: bacteremic pneumococcal pneumonia versus Mycoplasma pneumoniae pneumonia. Infection 25:90-94.[CrossRef][Medline] |
| 31. | Liechty, K. W., T. M. Crombleholme, D. L. Cass, B. Martin, and N. S. Adzick. 1998. Diminished interleukin-8 (IL-8) production in the fetal wound healing response. J. Surg. Res. 77:80-84.[CrossRef][Medline] |
| 32. | Lorian, V. 1986. Antibiotics in laboratory medicine. Williams & Wilkins, Baltimore, Md. |
| 33. | Maeyama, T., K. Kuwano, M. Kawasaki, R. Kunitake, N. Hagimoto, T. Matsuba, M. Yoshimi, I. Inoshima, K. Yoshida, and N. Hara. 2001. Upregulation of Fas-signalling molecules in lung epithelial cells from patients with idiopathic pulmonary fibrosis. Eur. Respir. J. 17:180-189. |
| 34. | Maniloff, J., R. N. McElhaney, L. R. Finch, and J. B. Baseman (ed.). 1992. Mycoplasmas: molecular biology and pathogenesis. American Society of Microbiology, Washington, D.C. |
| 35. | Matsumoto, M., M. Nishiguchi, S. Kikkawa, H. Nishimura, S. Nagasawa, and T. Seya. 1998. Structural and functional properties of complement-activating protein M161Ag, a Mycoplasma fermentans gene product that induces cytokine production by human monocytes. J. Biol. Chem. 273:12407-12414. |
| 36. | Nicholson, D. M., and N. A. Thornberry. 1997. Caspases: killer proteases. Trends Biochem. Sci. 22:299-306.[CrossRef][Medline] |
| 37. | Opitz, O., K. Pietsch, S. Ehlers, and E. Jacobs. 1996-1997. Cytokine gene expression in immune mice reinfected with Mycoplasma pneumoniae: the role of T cell subsets in aggravating the inflammatory response. Immunobiology 196:575-587.[Medline] |
| 38. | Pietsch, K., S. Ehlers, and E. Jacobs. 1994. Cytokine gene expression in the lungs of BALB/c mice during primary and secondary intranasal infection with Mycoplasma pneumoniae. Microbiology 140:2043-2048.[Abstract] |
| 39. | Rasmussen, S. J., L. Eckmann, A. J. Quayle, L. Shen, Y. Zhang, D. J. Anderson, J. Fierer, R. S. Stephens, and M. F. Kagnoff. 1997. Secretion of proinflammatory cytokines by epithelial cells in response to Chlamydia infection suggests a central role for epithelial cells in chlamydial pathogenesis. J. Clin. Investig. 99:77-87.[Medline] |
| 40. | Rawadi, G., and S. Roman-Roman. 1996. Mycoplasma membrane lipoproteins induce proinflammatory cytokines by a mechanism distinct from that of lipopolysaccharide. Infect. Immun. 64:637-643.[Abstract] |
| 41. | Rawadi, G., J. Zugaza, B. Lemercier, J. C. Marvaud, M. Popoff, J. Bertoglio, and S. Roman-Roman. 1999. Involvement of small GTPases in Mycoplasma fermentans membrane lipoproteins-mediated activation of macrophages. J. Biol. Chem. 274:30794-30798. |
| 42. | Razin, S. 1999. Adherence of pathogenic mycoplasmas to host cells. Biosci. Rep. 19:367-372.[CrossRef][Medline] |
| 43. | Rodwell, A. W., and R. F. Whitcomb. 1983. Methods for direct and indirect measurement of mycoplasma growth, p. 185-196. In S. Razin and J. G. Tully (ed.), Methods in mycoplasmology, vol. I. Mycoplasma characterization. Academic Press, New York, N.Y. |
| 44. | Schaffner, E., O. Opitz, K. Pietsch, G. Bauer, S. Ehlers, and E. Jacobs. 1998. Human pathogenic Mycoplasma species induced cytokine gene expression in Epstein-Barr virus (EBV)-positive lymphoblastoid cell lines. Microb. Pathog. 24:257-262.[CrossRef][Medline] |
| 45. | Stuart, P. M. 1993. Mycoplasmal induction of cytokine production and major histocompatibility complex expression. Clin. Infect. Dis. 17:S187-S191. |
| 46. | Thornberry, N. A., H. G. Bull, J. R. Calaycay, K. T. Chapman, A. D. Howard, M. J. Kostura, F. J. Miller, J. Chin, G. J. F. Ding, L. A. Egger, E. P. Gaffney, G. Limjuco, O. C. Palyha, S. M. Raju, A. M. Rolando, J. P. Salley, T.-T. Yamin, T. D. Lee, J. E. Shively, M. MacCross, R. A. Mumford, J. A. Schmidt, and M. J. Tocci. 1992. A novel heterodimeric cysteine protease is required for interleukin-1 beta processing in monocytes. Nature 356:768-774.[CrossRef][Medline] |
| 47. | Uchida, Y., H. Shiba, H. Komatsuzawa, T. Takemoto, M. Sakata, T. Fujita, H. Kawaguchi, M. Sugai, and H. Kurihara. 2001. Expression of IL-1beta and IL-8 by human gingival epithelial cells in response to Actinobacillus actinomycetemcomitans. Cytokine 14:152-161.[CrossRef][Medline] |
| 48. | Ulich, T. R., S. Yin, K. Guo, E. S. Yi, D. Remick, and J. del Castillo. 1991. Intratracheal injection of endotoxin and cytokines. II. Interleukin-6 and transforming growth factor beta inhibit acute inflammation. Am. J. Pathol. 138:1097-1101.[Abstract] |
| 49. | Wang, Y., and S. W. Walsh. 1996. TNF alpha concentrations and mRNA expression are increased in preeclamptic placentas. J. Reprod. Immunol. 32:157-169.[CrossRef][Medline] |
| 50. | Wingren, A. G., O. Bjorkdahl, T. Labuda, L. Bjork, U. Andersson, U. Gullberg, G. Hedlund, H. O. Sjogren, T. Kalland, B. Widegren, and M. Dohlsten. 1996. Fusion of a signal sequence to the interleukin-1 beta gene directs the protein from cytoplasmic accumulation to extracellular release. Cell. Immunol. 169:226-237.[CrossRef][Medline] |
| 51. | Wolfson, J. S., and D. C. Hooper. 1989. Fluoroquinolone antimicrobial agents. Clin. Microbiol. Rev. 2:378-424. |
| 52. | Yang, J., and P. J. Duerksen-Hughes. 2001. Activation of a p53-independent, sphingolipid-mediated cytolytic pathway in p53-negative mouse fibroblast cells treated with N-methyl-N-nitro-N-nitrosoguanidine. J. Biol. Chem. 276:27129-27135. |
| 53. | Yoshida, Y., M. Maruyama, T. Fujita, N. Arai, R. Hayashi, J. Araya, S. Matsui, N. Yamashita, E. Sugiyama, and M. Kobayashi. 1999. Reactive oxygen intermediates stimulate interleukin-6 production in human bronchial epithelial cells. Am. J. Physiol. 276:L900-L908. |
| 54. | Zhang, S., D. J. Wear, and S.-C. Lo. 2000. Mycoplasmal infections alter gene expression in cultured human prostatic and cervical epithelial cells. Immunol. Med. Microbiol. 27:43-50. |
This article has been cited by other articles:
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| J. Bacteriol. | J. Virol. | Eukaryot. Cell |
|---|
| Microbiol. Mol. Biol. Rev. | Clin. Vaccine Immunol. | All ASM Journals |
|---|
