Mutation in the transcriptional regulator PhoP contributes to avirulence of Mycobacterium tuberculosis H37Ra strain.

Attenuated strains of mycobacteria can be exploited to determine genes essential for their pathogenesis and persistence. To this goal, we sequenced the genome of H37Ra, an attenuated variant of Mycobacterium tuberculosis H37Rv strain. Comparison with H37Rv revealed three unique coding region polymorphisms. One polymorphism was located in the DNA-binding domain of the transcriptional regulator PhoP, causing the protein's diminished DNA-binding capacity. Temporal gene expression profiles showed that several genes with reduced expression in H37Ra were also repressed in an H37Rv phoP knockout strain. At later time points, genes of the dormancy regulon, typically expressed in a state of nonreplicating persistence, were upregulated in H37Ra. Complementation of H37Ra with H37Rv phoP partially restored its persistence in a murine macrophage infection model. Our approach demonstrates the feasibility of identifying minute but distinct differences between isogenic strains and illustrates the consequences of single point mutations on the survival stratagem of M. tuberculosisope

Comparison of virulence among mycobacterium tuberculosis isolates in Macrophages and in a latent infection model

의과학과/석사[한글]우리 나라 인구의 약 1/3 이상이 결핵에 감염되어 있는 것으로 추정되며, 매년 약 12만 명의 결핵환자가 발생한다고 보고되고 있다. 결핵균에 감염된 사람은 대부분 효과적인 면역 반응을 유발하여, 감염된 부위에 육아종을 형성하고, 질병의 진행을 차단한다. 임상 연구에 따르면, 육아종 내의 결핵균은 죽은 것이 아니라 잠복해 있다고 알려져 있다. 잠복 감염 상태의 대략 5-10%가 다시 재발하여 결과적으로 활동성이 있는 결핵을 유발한다.결핵의 잠복 감염에 영향을 주는 요인은 숙주의 면역 상태와 결핵균의 병원성 등으로 생각할 수 있다. 본 연구에서는 결핵 균주 간의 병원성의 차이가 잠복 감염 결핵균의 재활성화에 영향을 줄 것이라는 가설 하에 대식세포와 결핵균 잠복 감염 모델을 이용하여 한국에서 가장 많이 발견되는 결핵균 K균주와 다른 나라에서 분리된 임상 균주들 간의 병원성을 비교하고자 하였다.대식세포에서의 성장 곡선과 cytokine 생산을 비교한 결과 HN878, K, H37Rv, CDC1551 순으로 성장률이 낮았으며, K와 HN878로 감염된 대식세포에서 IL-6의 분비를 많이 유도하였다.결핵 잠복 감염 동물 모델에서 치료 3개월 후에 마우스 폐, 비장, 혈액에서 IFN-γ을 분비하는 세포 수를 균주 별로 비교한 결과 CDC1551이 숙주의 면역 반응을 가장 활발히 유도하였고, K는 숙주의 면역 반응을 가장 억제하는 양상을 보였다.요약을 하면, 각각의 결핵 균주에 따라 숙주는 다른 수준의 세포 매개 면역 반응을 유도하는 것으로 보인다. 또한 대식 세포 내에서의 결핵 균주의 증식과 감염성정도(infectivity)도 다른 것으로 보인다. 결핵 균주와 숙주간의 상호 작용을 규명하는 것은 앞으로 결핵에 대한 효과적인 백신과 치료 약제 개발에 도움이 될 것이라고 생각한다. [영문]Tuberculosis (TB), caused by Mycobacterium tuberculosis, is one of the world’s major infectious diseases. M. tuberculosis is a facultative intracellular pathogen residing in macrophages of the hosts. There have been many strain variations observed in M. tuberculosis over the several continents. Some of these M. tuberculosis strains caused outbreaks in several endemic and non-endemic areas. The virulence of these M. tuberculosis strains seemed to be related to many factors such as hosts’ immune responses, exposure of hosts to environmental mycobacteria, and strain variations. However, it has not been well characterized how different M. tuberculosis strains respond to the protective immune system of hosts. In this study, therefore, virulence of four M. tuberculosis strains (H37Rv, CDC 1551, HN878 and K) was compared by examining their growth rates in macrophages, induction of cytokines in macrophages and cell mediated immune responses in mice, and relapse rate in a latent infection model.When mouse bone marrow-derived macrophages were infected with M. tuberculosis, the HN878 strain was phagocytized most efficiently by or infected the macrophages, followed by the K and H37Rv strains in order. The CDC1551 strain was least infectious to macrophages. Number of live bacilli of these strains maintained at the approximately same level as the initial infection doses for six days. This may indicate that the K strain is as virulent as the HN878 strain.IL-6, a pro-inflammatory cytokine, was detectable in the highest concentration in the culture supernatant of macrophages infected with the K strain of M. tuberculosis and in lower concentration in the macrophages infected with either H37Rv or CDC1551. This may reflect bacterial load in the macrophages rather than efficiency of induction of IL-6 by M. tuberculosis strains.Lastly, a modified Cornell latent tuberculosis model was established and employed to compare relapse rate among the M. tuberculosis strains. When examined at 12 weeks after the last day of treatment, two to three animals of five animals per group had re-growth of M. tuberculosis in the lungs. In the spleen, however, both the K and HN878 strains had relapse in three out of five animals while H37Rv and CDC1551 only one or two, indicating that there may be a slight difference in extent of dissemination or relapse rate between M. tuberculosis strains. Interestingly, the CDC1551 strain induced strongly IFN-γ response while the K strain elicited a weak IFN-γ response, which may explain higher relapse in the latent infection model.In summary, there was a clear difference in efficiency of entering macrophages and in relapse rate in a latent infection model of TB in mice among M. tuberculosis strains. Such difference may be due to their ability of inducing cell-mediated immune response in ex vivo and in vivo models. The pathogenesis of M. tuberculosis may be, therefore, affected by the host factors in addition to the bacterial virulence. Further characterization of the interaction between M. tuberculosis and the host will help to develop the effective vaccine and therapeutic drugs for TB.ope

Filmkritik zu Stanley Kubricks Eyes Wide Shut unter besonderer Berücksichtigung der Sehtheorie Jacques Lacans

In dieser Arbeit wird versucht, Stanley Kubricks Film Eyes Wide Shut anhand der Lacanschen Sehtheorie zu analysieren, um zum einen die Anwendungsmöglichkeit der Lacanschen Theorie auf die Filmkritik systematisch zu zeigen und um sich zum anderen mittels eines neuen Gesichtspunktes kritisch mit Eyes Wide Shut auseinander zu setzen. In dem Film Eyes Wide Shut werden der Blick und das Sehen thematisiert, und genau genommen nicht nur auf der Figurenebene, sondern in einem irritierenden und herausfordernden Maße auch für die Zuschauer. Nach Lacan entsteht die menschliche Begierde aus dem Blick des Anderen, der auf den Mangel im Subjekt hinweist, und somit wirkt das Sehen entscheidend auf die Gestaltung des menschlichen Subjekts. Die Analyse wird unter folgenden zentralen Thesen der Lacanschen Sehtheorie unternommen: 1. Das Subjekt sieht, um sein Begehren zu befriedigen. Aber dieses Sehen führt nicht zur Erfüllung. 2. Das Subjekt ist nicht nur das sehende Wesen, sondern auch das erblickte Wesen. 3. Der begehrende Blick ist böse und die Kunst zähmt diesen bösen Blick. 4. Der Blick bringt das Reale zur Erscheinung. Der Film Eyes Wide Shut verhindert ständig das lustvolle Sehen, indem er die Szene, in der der Zuschauer Objekt a als den Grund und Gegenstand des Begehrens suchend ansieht, immer wieder unterbricht und die Identifizierung des Zuschauers mit der Hauptperson meistens nicht ermöglicht. Im Film bleibt die subjektive Verarbeitung der Vorfälle durch die Hauptperson weitgehend offen. Dies verlangt dem Zuschauer eine eigene Bewertung, Einordnung und Auslegung des Geschauten ab. Der Film organisiert ein Sehfeld, das den Blick des Zuschauers anlockt, und im nächsten Moment blockiert. Ein Wechselspiel der Erweckung von Schaulust und deren Enttäuschung ist ein Strukturprinzip des gesamten Films. Der Moment des Entzugs eröffnet manchmal einen Blick eines Anderen, der auf den Zuschauer zurückweist. Das Wechselspiel ist eine zähmende Funktion des Films, der den begehrenden Blick niederzulegen veranlasst. Der Film weist durch den Blick eines Anderen auf das Reale hin, das das Objekt a als Nichts entlarvt. Dadurch wird der Zuschauer sich des Mangels des menschlichen Wesens und der symbolischen Ordnung gewahr und somit wird eine Selbstwahrnehmung, Selbstbetrachtung und darüberhinaus eine neue Selbstgestaltung ermöglicht

Comparative analysis of immune responses to mycobacterium abscessus infection and its antigens in two murine models

Mycobacterium abscessus has been identified as an emerging pulmonary pathogen in humans. Because little is known regarding immune responses elicited by M. abscessus or its antigens, immunological responses were studied in two murine models subjected to intravenous (high-dose or systemic infection) or pulmonary (low-dose or local infection) inoculation with M. abscessus ATCC 19977. An overall comparison between the two models showed similar patterns of bacterial survival and host immune responses. The colonization of M. abscessus was the highest at 5 days post-infection (dpi) and its elimination was positively correlated with cell-mediated immunity in both challenges. However, an inverse relationship was observed between progressive inflammation and mycobacterial colonization levels in mice infected with a high dose at 14 dpi. Regarding antigens, culture filtrate (CF) of M. abscessus strongly induced IFN-gamma secretion, whereas cellular extract (CE) antigen elicited strong antibody responses. The antibody response to M. abscessus antigens in mice subjected to low-dose infection increased when the cellular immune response decreased over 14 dpi. However, the antibody response for the high-dose infection increased promptly after the infection. In comparison of cytokine expression in lung homogenates after M. abscessus infection, Thl and Th2 cytokines increased simultaneously in the high-dose infection, whereas only cell-mediated immunity developed in the low-dose pulmonary infection. These findings not only enhance our understanding of the immune response to M. abscessus infection according to systemic or pulmonary infection, but may also aid in immunological diagnosis and vaccine development. M. abscessus, murine infection model, immune response, antigens, cytokines.ope