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Research Activities | |||
The initial aim of the Center is the development of research projects in the field of inherited kidney disorders. In the future and depending on funding, there will be expansion in the direction of other clinical entities, in collaboration with other colleagues. The material collected and archived in the Biobank is typically DNA (from peripheral blood, or saliva, or other sources), plasma, serum, urine and a biopsy (renal or other, when available), along with demographic and medical information of the participating volunteers. All participants are informed by one of the medical doctors or other qualified research personnel and sign an informed consent form which has been approved by the Cyprus National Bioethics Committee (document EEBK3). All medical records and biological material are kept under secure conditions at a specially prepared Biobank space with absolute confidentiality and protection of sensitive data of personal character. Access to the data requires a password which is provided by the Coordinator to authorized personnel and collaborators. Similarly, access to the biological material is under the approval of the Coordinator and the Academic Council of the Research Center. There are several research directions in progress, combining molecular and cell biology approaches, clinical and molecular genetics, while a major research activity pertains to the characterization of a mouse model for Alport Syndrome. A general overarching aim is the understanding of the molecular and cellular processes that cause disease, thereby hoping to achieve better diagnosis, prevention and therapy of disease.
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Collaborating Centers / Partners | |||
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Brief description of projects | |||
1. Clinical and molecular investigation of familial microscopic hematuria of glomerular origin
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a) Nephropathies of Collagen Type IV | |||
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b. C3 Glomerulopathy (CFHR5 nephropathy) | |||
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2. Search for genetic modifiers that predispose patients with thin basement membrane nephropathy to focal segmental glomerulosclerosis and renal failure, on long follow-up. Also, follow-up of patients with CFHR5 nephropathy with the aim to search for genetic modifiers. | |||
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3. Follow up of patients with MUC1 kidney disease with the aim to search for early biomarkers and genetic modifiers | |||
Medullary cystic kidney disease (recently renamed MUC1 kidney disease), was diagnosed in several large families in Pafos, south-west of the island, in the 90s. MMRC researchers developed a clinical and genetics project that resulted in mapping the first gene on chromosome 1q21, in 1998 (Stavrou C et al, 1998;2002;Christodoulou K et al, 1998). It took several years before the gene was cloned by a Broad Institute group in the USA (Kirby A et al, 2013; Bleyer AJ et al, 2014). This development revived the interest of researchers and the Broad Institute group proposed a collaboration with MMRC in completing and upgrading the patient registry in Cyprus, which is presently in progress. It turns out that there are about 300 patients followed up in several counties, 150 of which are Greek-Cypriots, archived in our Biobank. Our common interests include the identification of potential early biomarkers, in blood or urine, for timely detection of the patients who run the highest risk to reach ESRD, usually at ages of 50s-70s. The gene is a very complex one and the mutation, a C insertion within a repeated region, is difficult to detect with traditional methods. Therefore, within the framework of this collaboration, all patients, in addition to linkage analysis at MMRC, are diagnosed with a proprietary certified method at Broad Institute.
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4. Investigation of the potential role of mir-548c-5p as a regulator of FOXC2 transcription | |||
Podocytes are highly differentiated epithelial cells outlining the glomerular vessels. FOXC2 is a transcription factor essential for inducing podocyte differentiation, development and maturation. It is considered to be the earliest podocyte marker and has been found to regulate the transcription of podocyte related genes. Murine Foxc2 knock-out podocytes fail to differentiate. In addition miRNAs are quite important in podocyte function as well, as podocyte specific dicer knockouts fail to maintain glomerular function. We hypothesize that the transcription of FOXC2 and consequently the differentiation of podocytes, can be influenced by miR-548c-5p through a predicted 21nt-long target region located 8kb upstream the FOXC2 transcription start point.
By using luciferase reporter constructs, it became apparent that the DNA target site acts as a conventional miRNA-binding site. During early differentiation of AB8/13 human podocytes, miR-548c-5p mimics effectively diminished endogenous FOXC2 levels, while a more dynamic model of this interaction was observed when investigating other time points, suggesting a dependence on target site availability. Therefore, the role of this miRNA target site as a distal enhancer/repressor element was investigated using Chromosome Conformation Capture. Evidence indicates a correlation between podocyte differentiation events and the interaction levels between the miRNA target site and the FOXC2 proximal promoter regions. Furthermore, human AB8/13 podocytes were transfected with miR-548c-5p mimics or inhibitors and FOXC2 transcription and translation levels were recorded. In addition, translation levels of WT1, another important podocyte transcription factor, were also determined. Neither of these two genes has a 3'UTR target site for miR-548c-5p. Cellular localization of FOXC2 levels was defined in tested differentiation time points by confocal microscopy. Collectively, results indicate an important coordination pattern between miR-548c-5p and FOXC2 transcription rates. This study can provide insights on the basic biology in regards to the initiation of podocyte differentiation and expand our understanding in podocyte phenotypes observed in glomerular disease. |
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5. Follow-up and characterization of a knock-in mouse model for Alport Syndrome, carrying mutation Col4a3-p.G1332E | |||
The mutation carried by the mouse is the recapitulation of the corresponding in human patients, COL4A3-p.G1334E, the most frequent founder mutation among Greek-Cypriots. At the moment 200 patients belonging to 18 families have been archived in our Biobank. Homozygous mice demonstrate activation of the unfolded protein response and have the characteristic ultrastructural pathology of Alport, with alternate thinning and thickening of the glomerular basement membrane on electron microscopy (Pieri M et al, 2014). We are in the process of completing the biochemical and other histological characterization of the mice, which have a rather mild phenotype overall. It is in our plans to use these mice for pre-clinical studies, treating them with synthetic chaperones, in the hope that relieving the ER stress will ameliorate the phenotype.
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6. Adolescent psychopathology cohort (this is a project developed and coordinated By Dr Kostas Fanti, Assistant Professor at the Department of Psychology of the University of Cyprus) | |||
Genetic risk for Antisocial Behaviour in adolescence-A Family-Based Association Study: This is a study of adolescents investigating the association of psychopathological symptoms in relation to distinct genetic risk markers. Specifically, saliva samples were collected from 210 adolescents and their parents, from which genomic DNA was extracted. In this sample, a complex disease paradigm is currently being employed to investigate the association between Conduct Disorder and multiple genes that code for a diverse array of neurobiological systems. These at risk adolescents were identified from a larger screening sample of 1,200 adolescents and their families that completed neuropsychological and behavioural measures of psychopathological symptoms, endophenotypes and personality traits. The aim in current and future work is to identify differential physiological and brain responses to emotional tasks to eventually propose a shift away from traditional diagnosis and a focus on core psychological processes that have a neurophysiological basis. Currently the researchers are studying gene by environment interactions to understand the development of behavioural problems. In addition, they are designing a study to investigate gene by neurophysiology interactions to further understand the development and aetiology of antisocial behaviours. Relevant Publications: Lordos, A. & Fanti, K. (under review). Executive Dysfunction and Callous Unemotional Traits mediate genetic risk for Developmental psychopathology lab: The Developmental Psychopathology lab was established in the Department of Psychology at the University of Cyprus by Dr Kostas A. Fanti. Following the Developmental Psychopathology perspective, the researchers combine questions about developmental change and psychopathology. Their research is primarily focused on investigating the development of different psychopathological problems, such as attention deficit hyperactivity disorder and conduct disorder, from preschool to adolescence. |