Ayşe Yüzbaşıoğlu

Assessment of housekeeping genes for use in normalization of real time PCR in skeletal muscle with chronic degenerative changes

The real time PCR technique requires the normalization of the gene of interest to reference genes that are accepted to be ubiquitously expressed. The choice of the reference gene(s) needs to be determined by researchers according to the particular tissue or model of interest. The best normalization gene is not easy to decide, particularly if the investigated tissue displays architectural changes and structural reorganization. We have investigated the expression of four housekeeping genes that are widely used for the normalization purposes (TATA binding protein, beta actin, hypoxanthine-guanine phosphorybosyl transferase and glyceraldehyde-3-phosphate dehydrogenase) in a skeletal muscle degeneration model applied by the release of the Achilles tendon which leads to a time-course degeneration of the soleus and gastrocnemius muscles. This study indicates that the TATA binding protein and the beta actin gene to be the least effected in the course of degeneration induced by tenotomy in rat soleus and gastrocnemius muscle.

Molecular analysis of Chanarin-Dorfman syndrome (CDS) patients: Identification of novel mutations in the ABHD5 gene

Chanarin-Dorfman syndrome (CDS) is an autosomal recessive metabolic disorder associated with congenital ichthyosis and a multisystemic accumulation of neutral lipids in various types of cells. Recently, mutations of the ABHD5 gene were identified as the cause of CDS. In this work, we carried out molecular analysis of the ABHD5 gene in 6 unrelated patients. We identified one previously reported mutation, N209X and two novel genetic alterations; a nonsense mutation (p.Y50X) and missense mutation (p.S73A).

Rationale, design and objectives of ARegPKD, a European ARPKD registry study.

BackgroundAutosomal recessive polycystic kidney disease (ARPKD) is a rare but frequently severe disorder that is typically characterized by cystic kidneys and congenital hepatic fibrosis but displays pronounced phenotypic heterogeneity. ARPKD is among the most important causes for pediatric end stage renal disease and a leading reason for liver-, kidney- or combined liver kidney transplantation in childhood. The underlying pathophysiology, the mechanisms resulting in the observed clinical heterogeneity and the long-term clinical evolution of patients remain poorly understood. Current treatment approaches continue to be largely symptomatic and opinion-based even in most-advanced medical centers. While large clinical trials for the frequent and mostly adult onset autosomal dominant polycystic kidney diseases have recently been conducted, therapeutic initiatives for ARPKD are facing the challenge of small and clinically variable cohorts for which reliable end points are hard to establish.Methods/DesignARegPKD is an international, mostly European, observational study to deeply phenotype ARPKD patients in a pro- and retrospective fashion. This registry study is conducted with the support of the German Society for Pediatric Nephrology (GPN) and the European Study Consortium for Chronic Kidney Disorders Affecting Pediatric Patients (ESCAPE Network). ARegPKD clinically characterizes long-term ARPKD courses by a web-based approach that uses detailed basic data questionnaires in combination with yearly follow-up visits. Clinical data collection is accompanied by associated biobanking and reference histology, thus setting roots for future translational research.DiscussionThe novel registry study ARegPKD aims to characterize miscellaneous subcohorts and to compare the applied treatment options in a large cohort of deeply characterized patients. ARegPKD will thus provide evidence base for clinical treatment decisions and contribute to the pathophysiological understanding of this severe inherited disorder.

Chanarin-Dorfman syndrome: Genotype-Phenotype Correlation

Chanarin-Dorfman syndrome is an autosomal recessive lipid storage disease characterized by non-bullous congenital ichthyosiform erythroderma, and involvement of the liver, muscles and central nervous system due to a multisystemic accumulation of neutral lipids in various types of cells. Less than 100 affected individuals have been reported worldwide, the majority from the Mediterranean and Middle-East countries, especially Turkey. We present clinical and molecular data of four affected relatives with Chanarin-Dorfman syndrome homozygous for a N209X mutation in ABHD5, and provide a short review by comparing patients with N209X homozygous mutations to patients with other ABHD5 mutations. No major clinical differences exist between individuals with an N209X mutation and those with other mutations, which argues against a genotype/phenotype correlation.

Changes in the Expression of Selenoproteins in Mesial Temporal Lobe Epilepsy Patients

Selenoproteins are enzymes containing selenium in their structure and are involved in cellular processes such as defense against oxidative stress and cell survival. The aim of this study is to investigate the expression of four selenoproteins (GPX1, TRXR1, SELP and SELW) in the hippocampus of intractable mesial temporal lobe epilepsy (MTLE) patients who underwent curative surgery. The selenoproteins is investigated at the mRNA level via RT-PCR and in situ hybridization and by immunostaining at the protein level. The expression of SELW exhibited a relative induction of more than tenfold, and immunostaining findings provided evidence that this upregulation is confined to neurons. GPX1 was also upregulated 2.3-fold, and TRXR1 was downregulated between 70 and 20% in MTLE patients. The profound induction of SELW has been accompanied by GPX1 and displayed a strong correlation with BCL2 expression, suggesting a protective role for these selenoproteins, and may be an indicator of a defense mechanism in surviving neurons.

Novel mutations of the MLC1 gene in Turkish patients.

Megalencephalic Leukoencephalopathy with Subcortical Cysts (MLC) is a rare autosomal recessive disease presenting with increased head circumference at birth or in early infancy. MLC1 (MIM 605908) mutations are responsible for this disorder. In this study, we sequenced the entire coding region of the MLC1 gene in 13 patients and detected five novel nucleotide variations in six of them. Two of the novel variations created a missense amino acid change and the other three were located in the introns and were putative splice mutations. One novel missense variation was observed in two unrelated patients from the central Black Sea region, and the data suggested a founder haplotype for this novel variation. Similarly, three unrelated patients with the previously reported p.Thr118Arg mutation shared a common haplotype. These data suggest an Anatolian origin for these two mutations. As in the previous reports, it is not possible to correlate the clinical phenotype of the patients with the mutation spectra.

Discovery of biomarkers in rare diseases: innovative approaches by predictive and personalized medicine

There are more than 8000 rare diseases (RDs) that affect >5 % of the world’s population. Many of the RDs have no effective treatment and lack of knowledge creates delayed diagnosis making management difficult. The emerging concept of the personalized medicine allows for early screening, diagnosis, and individualized treatment of human diseases. In this context, the discovery of biomarkers in RDs will be of prime importance to enable timely prevention and effective treatment. Since 80 % of RDs are of genetic origin, identification of new genes and causative mutations become valuable biomarkers. Furthermore, dynamic markers such as expressed genes, metabolites, and proteins are also very important to follow prognosis and response the therapy. Recent advances in omics technologies and their use in combination can define pathophysiological pathways that can be drug targets. Biomarker discovery and their use in diagnosis in RDs is a major pillar in RD research.

Chanarin-dorfman syndrome with multi-system involvement in two siblings.

Chanarin-Dorfman syndrome (CDS) is a very rare autosomal recessive inherited neutral lipid metabolism disorder associated with congenital ichthyosis and multi-system involvement. Observation of lipid vacuoles in neutrophils (Jordan’s anomaly) in peripheral blood smears in patients with ichthyosiform erythroderma is diagnostic. Herein we present 2 siblings with CDS that were referred to Dokuz Eylul University School of Medicine Department of Pediatrics due to ichthyosis. They had hepatomegaly, cataract, growth retardation, and sensorineural hearing loss. Some lipid vacuoles in neutrophils were noted in peripheral blood smear evaluation. Genetic analysis showed homozygous N209X mutation in both patients. They were put on a low-fat high-carbohydrate diet supplemented with medium-chain fatty acids. During 6 months of follow-up, no improvement was observed in both patients. In conclusion, although CDS is a rare lipid storage disease, it should always be a consideration in patients with congenital ichthyosis, especially those with extracutaneous symptoms or signs. The diagnosis of CDS is made based on a very simple test-peripheral blood smear. Conflict of interest:None declared.

Biobanks as Basis of Individualized Medicine: Challenges Toward Harmonization

Abstract Biobanking is a recent field where human biospecimens and related personal and clinical data are collected and stored for long-term use in biomedical research. The governance models for biobanking research need to involve all stakeholders and must be organized in both ethics and legal aspects and quality management systems. Today, biobanking is moving toward a global scale, and harmonization of all governance steps must be well defined to ensure that a larger research community can benefit from shared materials of high standards. Main ethical issues involved are mostly around protection of donor privacy, informed consent where individual autonomy must be exercised should be a norm, and use of harmonized consent model can facilitate international networking. Quality management of biobanks assures high quality and security of biosamples and associated data; this further fosters public trust and utility of sample collections. This chapter summarizes current governance structures and challenges for international harmonization of biobanks.