Chlamydia (Chlamydophila) pneumoniae is a common respiratory pathogen, and it seems likely that alveolar macrophages may have an important role in infection with this bacterium. In the present study, we examined the usefulness of a continuous cell line of murine alveolar macrophages, designated "MH-S," as an in vitro C. pneumoniae infection model. Infection of MH-S cells with C. pneumoniae resulted in the development of typical inclusion bodies in the cells, similar to that seen in primary alveolar macrophages. However, we noted that, although the number of bacteria in the cultures increased during the infection, there was a restricted production of infective elementary bodies. The analysis of bacterial messenger RNA in the cultures showed that the message levels for the omcB gene were present only at a moderate level, but the levels of hsp60 messages increased markedly during infection. Neutralization of tumor necrosis factor (TNF)-alpha induced by inoculation with antibody significantly enhanced the infection, but omcB message levels were still inhibited. These results indicate that the growth of C. pneumoniae in alveolar macrophages may be restricted. Endogenous TNF-alpha may be one of the factors responsible for such restriction, but other factors also may be involved.

Since current studies indicate the possible involvement of Chlamydia pneumoniae in the pathogenesis of multiple sclerosis (MS), demonstration of C. pneumoniae in the cerebrospinal fluid (CSF) of patients with MS is highly desirable. However, there is controversy concerning the detection of C. pneumoniae in CSFs from MS patients due to the lack of a standard protocol for extraction and detection of C. pneumoniae DNA. In this regard, we attempted to establish a highly effective extraction protocol for C. pneumoniae DNA from CSFs utilizing a commercial kit and a PCR detection method. The extraction and PCR detection protocol established in this study succeeded in detecting as few as 20 C. pneumoniae organisms in 200 l of mock CSF. The use of this protocol to detect C. pneumoniae DNA in CSFs revealed that 68% of CSF samples obtained from patients with MS were positive (11 out of 16 samples) for chlamydia DNA. Thus, the protocol established here is sensitive enough to detect chlamydia DNA from CSFs and can be used by other laboratories for evaluation of the presence of chlamydiae in CSFs because the protocol is based on the use of a commercial kit.Multiple sclerosis (MS) is a chronic demyelinating disease of the central nervous system (CNS) characterized by focal areas of demyelination. Although the exact etiology of MS is unknown, it is generally accepted that autoimmunity is involved and that the autoantigen(s) probably resides in CNS myelin, the target of the immune response (1). In this regard, current studies argue for an infectious agent as an initiating or enhancing factor for MS with immunological mechanisms (5). To identify a specific causative agent for MS, many groups have attempted to detect microbes in cerebrospinal fluid (CSF) as well as in CNS lesions obtained from MS patients. However, no consistent results have been obtained with any given pathogen. Recent studies conducted by Sriram et al. (12) highlighted the possible involvement of a bacterium in MS, with the finding of Chlamydia pneumoniae in the CSF of almost all patients with MS but in only a small proportion of CSF samples from control subjects without MS. That study has shown the highest association of any organism with MS to date. However, other research groups either could not detect C. pneumoniae in CSFs from MS patients or detected it only in a small proportion of specimens (2,8,14). This may be due to the lack of a standard method for C. pneumoniae detection in CSFs. For study of the involvement of C. pneumoniae in the pathogenesis of MS, a reliable standard evaluation protocol for C. pneumoniae in clinical specimens is essential. Therefore, in the present study, we attempted to establish an efficient extraction protocol for C. pneumoniae DNA in CSFs by use of a commercial kit followed by PCR specific for C. pneumoniae. Furthermore, the extraction and detection system established for C. pneumoniae DNA was applied to demonstration of the presence of C. pneumoniae in CSFs obtained from patients with MS. The results indicate that the proto...

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