Pozzobon, Tommaso
Angiogenesis in secondary syphilis: role of the bacterioferritin TpF1, antigen of Treponema pallidum [Tesi di dottorato]

Syphilis is a chronic inflammatory disease transmitted by the spirochete Treponema pallidum. The bacterium is usually acquired by direct contact, generally sexual; the disease is worldwide spread and it represents an important global health problem. Syphilis is indeed a major sexually transmitted disease, particularly in developing countries, but an increasing number of new cases have also been recently noted in USA and Europe. Syphilis is a progressive and multistage disease with diverse and wide-ranging manifestations. Syphilis is an evolving disease and can be divided in three main stages. In the first stage, a red papule appears at the site of inoculation, usually genitals. Within the first few days, the papule ulcerates, producing the typical chancre of primary syphilis, which is painless and filled with treponemes. The chancre is accompanied by regional lymphadenopathy. If untreated, bacteria can also proliferate in the chancre and can be transported via lymphatics to the bloodstream, from which they disseminate throughout the body, giving rise to secondary syphilis: this represents the second stage and is a systemic disease. The most common manifestation of secondary syphilis is a disseminated mucocutaneous rash, which usually resolves spontaneously. In untreated patients, the symptoms are absent for a variable period of time, which is called latent stage. Afterwards, one-third of untreated patients can develop symptomatic late syphilis: the third stage of the disease, which is subdivided into gummatous syphilis, neurosyphilis and cardiovascular syphilis. This disease could be cured with an antibiotic treatment, but, if not adequately treated, syphilis can lead patients to death. Nowadays there are no available vaccines to prevent syphilis in particular due to the difficulties in cultivating T. pallidum in vitro make more difficult to study this pathogen as well as the identification of its virulence factors. Little is known about the mechanism by which the bacterium causes the clinical manifestations that accompany the different stages of the disease, however it is established that T. pallidum is able to invade and survive in a wide variety of tissues and organs and that it promotes the formation of new blood vessels. Angiogenesis could have a crucial role in syphilis pathogenesis, at least for two reasons: i) the bacterium has limited metabolic capabilitie, thus probably it requires support from the host and it may derive most essential macromolecules from the blood. ii) Increased vascular permeability is one of the first stages in angiogenesis and the organism could take advantage of the vascular leakage to access to and egress from the bloodstream, resulting in systemic spread. Another distinctive feature of syphilis consists in the fact that it becomes chronic in untreated patients and this probably reflects the ability of the bacterium in eliciting a T regulatory response, which could be associated with the fading of the host effector immune responses against the pathogen. We have recently demonstrated that the bacterioferritin TpF1, a major antigen of T. pallidum, plays a pivotal role in driving this suppressive immune response, by modulating the release of specific cytokines by monocytes. TpF1 is a protein homolog to another immunomodulant antigen produced by the bacterium Helicobacter pylori and called HP-NAP. Both these proteins belong to the Dps-like family, a versatile group of bacterial stress miniferritins with a nearly spherical dodecameric structure. HP-NAP, besides interacting with neutrophils, monocytes and dendritic cells and modulating their activity (as TpF1 does), also binds to endothelial cells, in which it is internalized by transcytosis. Considering the homology between HP-NAP and TpF1 and the fact that angiogenesis is a characteristic of the secondary syphilis, we wondered whether TpF1 could interact with endothelial cells and promote their proliferation. We have demonstrated that TpF1 induces proliferation and migration of human endothelial cells (HUVECs); moreover, the protein activates endothelial cells to form microcapillary-like structures in vitro inducing marked changes in the cell pattern with the formation of tubules assembled by elongation and joining of the cells; a similar pattern was observed with VEGF. With the aim of understanding the mechanism underlying the activity of TpF1, we have addressed the possibility of an indirect effect mediated by VEGF or IL-8, both proangiogenic factors. We have excluded any contribution by VEGF: indeed, the latter is not released by the endothelial cells upon TpF1 stimulation. On the contrary, we have found that TpF1 triggers the release of IL-8 and, notably, the blockage of the cytokine strongly prevents the angiogenic activity of the bacterial protein inhibiting endothelial cell proliferation, migration and micro capillary-like structures formation. We also verified that the secretion of IL-8 relies on the induction of intracellular cAMP, produced by cellular adenylate cyclases, which is able to induce gene transcription through the activation of the two transcription factors NF-?B and CREB (cAMP response element-binding protein). Finally we have observed that TpF1 induces new blood vessels formation in vivo in a zebrafish model. Moreover TpF1 promotes, in zebrafish, IL-8 gene expression suggesting a role for this cytokine in the TpF1-induced angiogenesis process also in vivo.

In relazione con http://paduaresearch.cab.unipd.it/7919/
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BIO/13 - Biologia applicata


Tesi di dottorato. | Lingua: | Paese: | BID: TD16055131