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Familial Mediterranean Fever

#A-1055


The Role of Intestinal Microflora on the Pathogenesis Familial Mediterranean Fever

Tech Area / Field

  • BIO-CGM/Cytology, Genetics and Molecular Biology/Biotechnology
  • BIO-CHM/Biochemistry/Biotechnology
  • MED-OTH/Other/Medicine
  • BIO-MIB/Microbiology/Biotechnology

Status
8 Project completed

Registration date
21.07.2003

Completion date
05.11.2008

Senior Project Manager
Visser H

Leading Institute
Institute of Molecular Biology, Armenia, Yerevan

Collaborators

  • University of Aberdeen / Department of Medicine and Therapeutics, UK, Aberdeen\nRowett Research Institute, UK, Aberdeen\nUniversity of Dundee / Ninewells Hospital Medical School, UK, Dundee

Project summary

1. The goal of the project. The main goal of this project is to understand the role of gut bacteria (direct or indirect via metabolites) in the etiopathogenesis of FMF. Using molecular analysis of intestinal microflora of FMF patients, we will establish which bacterial groups could be responsible for the onset and progression of the disease. These groups will be isolated and tested for the induction of immune response (pro- and anti-inflammatory pathways). This basic research will lay a foundation for the subsequent applications such as designation of alternative strategies to prolong the period of remission and alleviate the severity of disease by using correcting diet, probiotic, prebiotic, and synbiotic therapies.

2. The state of the art in the field. Familial Mediterranean fever has become recognized as an independent nosological unit after the discovery of the chromosomal defect inherited in this disease. But still, FMF remains a disease, which etiopathogenesis is far from being clear yet. The tendency toward the increased morbidity rate and the broadening spectrum of the ethnic base for FMF, which has been earlier observed only within the Mediterranean populations (Armenians, Jews, and Arabs), gives the basis to reconsider the opinion that FMF is ethnically stipulated. In this regard, elucidation of the etiopathogenesis of this disease becomes extremely important. Despite that disease has been known for several millennia, the MEFV gene responsible for it (or, more correctly, its mutated alleles) has been positionally cloned by two independent consortia only in 1997. The following investigations placed the gene in the myelomonocytic-specific pro-inflammatory pathway and identified it as an IFN-gamma immediate early gene. FMF is a recessive disorder characterized by episodes of fever and neutrophil-mediated serosal inflammation. On the other side, geno-phenotypical analyses suggest a considerable involvement of environmental modifier-factors such as change of diet, stress, and other unidentified factors. Moreover, periodic nature of this disease is certainly indicative of environmental factors. While major advances were made towards understanding genetics of this disease, its modulation by environmental factors is poorly understood. In this proposal, we will investigate the role of intestinal bacteria in the development of this disease. Human gut microflora plays an important role in shaping and maintaining the normal mucosal immunity through the production of short chain fatty acids and proper antigenic signaling to the host. There is a delicate equilibrium between the gut microflora and the host’s immune system, which allows to maintain a proper balance of pro- and anti-inflammatory responses. We hypothesize that in FMF patients this balance is disturbed because of the changed bacterial composition of the gut. This biased bacterial population sends “wrong” signals to the host thus launching a cascade of inflammatory reactions leading to clinical manifestations such as abdominal pain, fever, and serosal inflammation. Presently, however, the role of intestinal bacteria in this process is completely unknown. Our preliminary investigations suggest that changes in the intestinal microflora can trigger severe disease attacks in FMF patients. In particular, our data demonstrated that the blood of FMF patients has the cardinally biased and unusual profile of small molecules of bacterial origin implying that the gut microflora of these patients is extremely deviant from the norm. The abnormal microflora, in turn, may aggravate the patient’s conditions through the production of abnormal antigens and small molecules of bacterial origin. Notably, such type of micro-ecological breakage, which affects the human organism, has not been described yet.

3. The impact of the proposed project on the progress in this field. The concept of distorted microflora in FMF patients, which leads to imbalance in microbe-host interaction and serosal inflammation process, is a pioneering effort toward understanding of FMF pathology. This investigation will provide valuable information on the way gut microorganisms interact with the host’s immune system. It will contribute to further understanding of mechanisms of various gut pathologies including cancerous growth. Ultimately, this knowledge would help to design the strategies to down-regulate the abnormal inflammatory responses by taking specific food additives such as pro- and pre-biotics, changing the diet as well as targeting specific groups of bacteria by narrow-range antibiotics. In the light of tendencies toward the increased morbidity rate and the broadening spectrum of the ethnic base for FMF, the results of this project may have broader impact exceeding the previously recognized ethnic base.

4. The competence of the project team in the specified area. Presently, our scientific group is working under some aspects of the project such as determination of small molecules of bacterial origin in the blood of FMF patients. However, to study the problem in its entity additional funding is required. Despite the shortage of funding, the group has already made a significant contribution to this problems, which is reflected in our publications. The group consists of highly-qualified specialists in the fields of molecular biology, molecular microbiology, immunology, biochemistry, and also clinical physicians dealing with FMF. The group is the only one in the Republic of Armenia, which carries out works on a high scientific-technical level, using modern molecular technologies equivalent to international standards.

The scientific leader of the project – Acad. Karageuzyan K.G. is a recognized scientist with a wide range of professional interests including various aspects of medical and biological sciences. The results of his pioneering research on studying the metabolic disturbances of phospho- and glyco-lipids have been presented in more than 500 articles. This enormous experience is certainly helpful for understanding the mechanisms of initiation, development, and generalization of pathologies of different origins. The manager of the project Sarkisyan N.N. - Ph.D., has more than 30 years of experience in the field of molecular biology and biochemistry of microorganisms, familiar with modern methods used in microbial ecology of health and disease, has more than 45 scientific publications. Our team also includes the investigators from Moscow (Russia), which are the authors of the original SMOM homeostasis concept and which have participated in obtaining the preliminary results. The collaborators of this project are Dr. Aminov R. (UK), Dr. Kelly D. (UK), Dr. Aminova L. (USA), Dr. Takenaka A. (Japan).

5. Expected results. Proposed research project encompasses both basic and applied research. This is a first complex study of its kind, which will include analysis of genetic background of patients, their immune status, circulation of bacterial metabolites in the blood, and analysis of gut microflora. First, with the use of 16S rDNA libraries, we will establish, what kind of the microflora bias is common for FMF patients in comparison with the normal state. This would allow to identify groups of bacteria that are responsible for clinical manifestation of the disease. Second, we will isolate these bacteria and characterize them for production of specific metabolites, namely SMOM and SCFA. This part will answer the question, what kind of microbially produced substances could be harmful for the host. Third, we will analyze the genetic background and immune status of the same patients. Based on these analyses we will create a database to correlate all parameters such as severity of disease, the presence of specific bacteria, production of specific metabolites, and genetic and immune status of patients. We expect that for the first time, this complex study will clarify the role of intestinal microflora in development, progression, remission, and acute phases of FMF. It will provide valuable information on host-microbe interaction and on what bacterial molecules are responsible for the signaling to the host. We also will identify diagnostically significant microbial markers specific for this disease. This basic knowledge will allow several potential applications. First, bring new recommendations on treatment and prophylaxis of FMF exacerbation, the strategies for the adequate complex therapy including the proven treatments such as colchicine to prolong the periods of remission. Second, development of bacterial diagnostics for early detection of the disease. We plan to publish several articles in international journals. The results will be also made public through newspapers, popular journals, Web pages, presentations in educational institutions, on radio and television.

6. Meeting ISTC Goals and Objectives.

1. The project is of great value and importance because the scientists of the Institute of Molecular Biology NAS RA, which previously were involved into defensive research in the former USSR, will have the possibility to work in civil medical area. 50% of the studies in this project will be carried out by these scientists.

2. The project also cooperates to the integration of the scientists from the states of FSU to the International Scientific Cooperation. In the project participate investigators from Research group of Academician Yu. Isakov, Russian Academy of Medical Science, Moscow, Russia: Prof. Beloborodova NV, Prof. Osipov GA.

3. The project is directed to the investigation and designation of technologies in the field of human microbial ecology and understanding the molecular mechanisms of gut microflora and host organism’s interaction, its role in human physiology and in the development of pathologic condition, particularly in FMF.

7. Scope of activities. The project is planned for three years. The basic research will be carried out on blood and fecal samples of 200 FMF patients (in attack and remission periods) and groups of comparison (100 healthy donors and 100 with other than FMF gastrointestinal diseases) for reaching statistically significant conclusions. At the same time, data set of clinical manifestation will be collected. The specific objectives of this project are: 1. Identification of gut microbiota alteration in FMF patients in comparison with normal microbiota; 2. Screening the blood for bacterial metabolites (SMOM and SCFA) and identification of prevalent markers in FMF patients; 3. Identification of microbial metabolites responsible for the launch of inflammatory processes in FMF; 4. Investigation of cytokine net in FMF patients (in attack and remission periods); 5. Establish the correlation between gut microflora bias and abnormalities of immune status; 6. Analysis of correlation between the SMOM and SCFA, taxonomic affiliation of microorganisms, immune status, and disease state in FMF patients.

On the basis of data obtained the recommendations on treatment and prophylaxis of FMFs’ exacerbation will be elaborated and presented to the Ministry of Health with the aim of their realization. These recommendations will be also directed to end users such as practicing clinicians. Implementation of the project objectives will be realized through the stages described in the project by all participants during three years.

8. The role of foreign collaborators. The cooperation with foreign collaborators is proposed: coordination between the investigators, as well as assessments of progress, will be primarily through the electronic mail. The other aspect of cooperation will be coordinated through the reciprocal visits. These visits will also allow data analyses, discussions, and preparation of results for reports and publication.

9. Technical approach and methodology. The methodology proposed for this work such as gas chromatography-mass spectrometry (GC-MS), immunoferment assay (IFA), microbial community analysis through sequencing of 16S rDNA libraries has the potentials to uncover the role of microorganisms in FMF etiopathogenesis. The method of GC-MS allows screening the large number of microbial markers and metabolites in one sample of clinical material. The series of investigations will be carried out during the FMFs’ exacerbation, in dynamics, and in the stage of remission with the same patient to ensure the diagnostic significance of markers. Community analysis through 16S rDNA sequencing allows taxonomical identification of bacteria suspected in the FMF pathogenesis. Estimation the degree of participation of mediators (C5a, pro- and anti-inflammatory cytokines) in immune response will be carried out using the immunoferment assay (IFA). This model of complex studies including molecules of bacterial origin, taxonomic affiliation of microorganisms, immune status, and the state of disease in FMF patients, will help to understand the role of the microflora deviations in FMF patients, which would help to design alternative strategies to alleviate the severity of disease by using corrective diet, probiotic supplements, synbiotic therapies, and antibiotic therapy to suppress bacteria involved in the pathogenesis of FMF.


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The International Science and Technology Center (ISTC) is an intergovernmental organization connecting scientists from Kazakhstan, Armenia, Tajikistan, Kyrgyzstan, and Georgia with their peers and research organizations in the EU, Japan, Republic of Korea, Norway and the United States.

 

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