UGT1A1 (TA)n Promoter Genotype: Diagnostic and Population Pharmacogenetic Marker in Serbia.

The UGT1A1 enzyme is involved in the metabolism of bilirubin and numerous medications. Unconjugated hyperbilirubinemia, commonly presented as Gilbert syndrome (GS), is a result of decreased activity of the UGT1A1 enzyme, variable number of TA repeats in the promoter of the UGT1A1 gene affects enzyme activity. Seven and eight TA repeats cause a decrease of UGT1A1 activity and risk GS alleles, while six TA repeats contribute to normal UGT1A1 activity and non-risk GS allele. Also, the UGT1A1 (TA)n promoter genotype is recognized as a clinically relevant pharmacogenetic marker. The aim of this study was to assess diagnostic value of UGT1A1 (TA)n promoter genotyping in pediatric GS patients. Correlation of the UGT1A1 (TA)n genotypes and level of unconjugated bilirubin at diagnosis and after hypocaloric and phenobarbitone tests in these patients was analyzed. Another aim of the study was to assess pharmacogenetic potential of UGT1A1 (TA)n variants in Serbia. Fifty-one pediatric GS patients and 100 healthy individuals were genotyped using different methodologies, polymerase chain reaction (PCR) followed by acrylamide electrophoresis, fragment length analysis and/or DNA sequencing. Concordance of the UGT1A1 (TA)n promoter risk GS genotypes with GS was found in 80.0% of patients. Therefore, UGT1A1 (TA)n promoter genotyping is not a reliable genetic test for GS, but it is useful for differential diagnosis of diseases associated with hyperbilirubinemia. Level of bilirubin in pediatric GS patients at diagnosis was UGT1A1 (TA)n promoter genotype-dependent. We found that the frequency of pharmacogenetic relevant UGT1A1 (TA)n promoter genotypes was 63.0%, pointing out that UGT1A1 (TA)n promoter genotyping could be recommended for preemptive pharmacogenetic testing in Serbia.

Genetic characterization of GSD I in Serbian population revealed unexpectedly high incidence of GSD Ib and 3 novel SLC37A4 variants.

Glycogen storage disease (GSD) type I is inborn metabolic disease characterized by accumulation of glycogen in multiple organs. We analyzed 38 patients with clinical suspicion of GSD I using Sanger and next-generation sequencing (NGS). We identified 28 GSD Ib and 5 GSD Ia patients. In 5 patients, GSD III, VI, IX, cholesteryl-ester storage disease and Shwachman-Diamond syndrome diagnoses were set using NGS. Incidences for GSD Ia and GSD Ib were estimated at 1:172 746 and 1:60 461 live-births, respectively. Two variants were identified in G6PC gene: c.247C>T (p.Arg83Cys) and c.518T>C (p.Leu173Pro). In SLC37A4 gene, 6 variants were detected. Three previously reported variants c.81T>A (p.Asn27Lys), c.162C>A (p.Ser54Arg) and c.1042_1043delCT (p.Leu348Valfs*53) accounted for 87% of all analyzed alleles. Computational, transcription studies and/or clinical presentation in patients confirmed pathogenic effect of 3 novel variants: c.248G>A (p.Gly83Glu), c.404G>A (p.Gly135Asp) and c.785G>A (p.Ser263Glyfs*33 or p.Gly262Asp). In the cohort, hepatomegaly, hypoglycemia and failure to thrive were the most frequent presenting signs of GSD Ia, while hepatomegaly and recurrent bacterial infections were clinical hallmarks of GSD Ib. All GSD Ib patients developed neutropenia while 20.6% developed inflammatory bowel disease. Our study revealed the highest worldwide incidence of GSD Ib. Furthermore, description of 3 novel variants will facilitate medical genetic practice.

Molecular genetic study of congenital adrenal hyperplasia in Serbia: novel p.Leu129Pro and p.Ser165Pro CYP21A2 gene mutations.

PURPOSE: Congenital adrenal hyperplasia (CAH) is an autosomal recessive disease characterized by impaired adrenal steroidogenesis and most often caused by CYP21A2 gene mutations. For the first time, we reported complete spectrum and frequency of CYP21A2 gene mutations in 61 unrelated patients with classical and non-classical CAH from Serbia. METHODS: Direct DNA sequencing of whole CYP21A2 gene and polymerase chain reaction with sequence-specific primers for detection of CYP21A1P/CYP21A2 chimeras were combined. RESULTS: We identified 18 different pathogenic alleles-two of them novel. Mutation detection rate was highest in patients with salt-wasting form of CAH (94.7%). The most prevalent mutation was intron 2 splice site mutation, c.290-13A/C>G (18.5%). Other mutation frequencies were: CYP21A1P/CYP21A2 chimeras (13%), p.P30L (13%), p.R356W (11.1%), p.G110fs (7.4%), p.Q318X (4.6%), p.V281L (4.6%), p.I172N (2.8%), p.L307fs (2.8%), p.P453S (1.9%), etc. Mainly, frequencies were similar to those in Slavic populations and bordering countries. However, we found 6.5% of alleles with multiple mutations, frequently including p.P453S. Effects of novel mutations, c.386T>C (p.Leu129Pro) and c.493T>C (p.Ser165Pro), were characterized in silico as deleterious. The effect of well-known mutations on Serbian patients' phenotype was as expected. CONCLUSIONS: The first comprehensive molecular genetic study of Serbian CAH patients revealed two novel CYP21A2 mutations. This study will enable genetic counseling in our population and contribute to better understanding of molecular landscape of CAH in Europe.

[Modern digital technologies as a useful tool for dental implantation planning].

The method of implant size choice concerning mechanical properties of bone tissue surrounding an implant is proposed based on CT-assessment of anatomical and topographical conditions using MIMICS-ANSYS software.

Hess Lancaster screen test with the head tilted: a useful test in the diagnosis of bilateral fourth nerve palsies.

PURPOSE: To introduce Hess Lancaster screen test performed with the head tilted as a new ancillary test to help in the diagnosis of bilateral superior oblique (SO) nerve paresis. METHODS: Three patients with clinical diagnosis of acquired bilateral fourth nerve paresis, three with clinical diagnosis of acquired unilateral fourth nerve paresis, and three normal subjects were tested with Hess Lancaster test in the nine positions of gaze with their heads straight and with their heads tilted to each side. Test results were compared and data were analyzed. RESULTS: Hess Lancaster test of all patients with bilateral SO palsy performed with their heads straight showed unilateral SO underaction, but when the test was carried out with the head tilted to each side findings were positive for ipsilateral SO underaction and ipsilateral hypertropia for both sides. In those patients with unilateral SO palsy, Lancaster test was positive for SO underaction when the head was in straight position and when it was tilted to the side of the paretic muscle, but it was almost normal with the head tilted to the opposite side. Normal subjects did not show any abnormality regardless of the head position. CONCLUSIONS: Bilateral SO palsies are sometimes difficult to diagnose for they might be masked. Hess Lancaster test carried out with the head tilted to both sides could help in the diagnosis of bilateral SO palsies.

Disinsertion of the medial rectus following pterygium surgery: signs and management.

The clinical characteristics, intraoperative findings and management of two cases of "lost" medial recti during surgery for recurrent pterygium are described. The lost muscles are classified in different groups according to their etiopathogenesis. In the cases reported, the muscles were found retroinserted with extensive proliferation of fibrous tissue. This complication was resolved after finding the muscle, liberating the surrounding fibrous tissue and reattachment in its original insertion. Two aspects stand out: The CAT scan to determine the location of the muscle and the usefulness of topical anesthesia to facilitate recognition of the muscle during the surgical procedure.