Congenital Proximal Radioulnar Synostosis in an Elite Athlete-Case Report.

Background and Objectives: Proximal radioulnar synostosis (PRUS) is the most frequent congenital forearm disorder, although the prevalence in the general population is rare with a few hundred cases reported. Pfeiffer, Poland, Holt-Oram, and other serious congenital syndromes contain this abnormality. Non-syndromic cases with isolated PRUS very often exhibit as SMAD6, NOG genes variants, or sex chromosome aneuploidy. A subgroup of patients with haematological abnormalities presents with HOXA11 or MECOM genes variants. Case report: We present a non-syndromic adult elite ice-hockey player with unilateral proximal radioulnar synostosis of the left forearm. In early childhood he was able to handle the hockey stick only as a right-handed player and the diagnosis was set later at the age of 8 years due to lack of supination. Cleary-Omer Type III PRUS was found on x-ray with radial head hypoplasia and mild osteophytic degenerative changes of humeroulnar joint. Since the condition had minimal impact on sports activities, surgical intervention was not considered. The player continued his ice-hockey career at the top level and joined a national team for top tournaments. Upper extremity function assessment with questionnaires and physical testing resulted in minimal impairment. The most compromised tool was the Failla score with 10 points from a total of 15. Genetic testing with Sanger sequencing revealed no significant pathogenic variant in SMAD6, NOG, and GDP5 genes. No potentially pathogenic copy number variants were detected by array-based comparative genomic hybridization. Conclusions: In the reported case, the ability of an athlete to deal with an anatomic variant limiting the forearm supination is demonstrated. Nowadays, a comprehensive approach to rule out more complex musculoskeletal impairment and family burden is made possible by evolving genetics.

Prevalence of Low Serum Alkaline Phosphatase and Hypophosphatasia in Adult Patients with Atypical Femur Fractures.

Hypophosphatasia (HPP) is a rare genetic disorder characterized by low serum alkaline phosphatase (ALP), its manifestations may include atypical femoral fractures (AFF). However, the prevalence of low serum ALP and HPP in patients with AFF remains unknown. We retrospectively analyzed ALP levels and clinical manifestations compatible with HPP in 72 adult patients with confirmed AFF by chart review. ALP values were compared with those of a control group of patients with prior proximal femoral fracture during antiresorptive treatment (n = 20). Among the AFF patients, 18 (25%) had at least one serum ALP value ≤ 40 IU/L, although in all but one case, at least one ALP value > 40 IU/L was also detected at another time point. Most low ALP values were associated with antiresorptive treatment (P = 0.049) and lowest levels of ALP did not differ between the AFF and the control groups (P = 0.129). However, low ALP values among AFF patients were associated with a higher rate of bilateral AFF (50% vs 22%, P = 0.025), metatarsal fracture (33% vs 7%, P = 0.006), and with trends for more frequent use of glucocorticoid (22% vs 8%, P = 0.089) and proton pump inhibitor (61% vs 44%, P = 0.220). In one AFF patient with low ALP and clinical suspicion of HPP, a rare pathogenic heterozygous variant of the ALPL gene was identified. In conclusion, low ALP values are common among subjects with AFF and mainly related to concomitant antiresorptive medication. Hence, low serum ALP has low specificity for HPP among AFF patients.

TREM2 coding variants in Slovak Alzheimer's disease patients.

BACKGROUND: Triggering receptor expressed on myeloid cells 2 (TREM2) is an important modulator of innate immune responses. In the human brain, TREM2 is primarily expressed on microglia and is involved in cell survival, phagocytosis, and regulation of inflammation. TREM2 dysfunction has been linked to the pathogenesis of various neurodegenerative diseases including Alzheimer's disease (AD). Rare coding variants of the TREM2 gene have been reported to modulate AD risk in several populations, however, data on their association with susceptibility to AD in the Slovak population have been missing. METHODS: We have analyzed 10 non-synonymous coding variants located in TREM2 exon 2 by direct sequencing in 270 late-onset Alzheimer's disease (LOAD) patients and 331 controls. RESULTS: Four out of 10 TREM2 mutant variants have been identified in the analyzed groups, namely rs75932628 C > T (R47H), rs142232675 C > T (D87N), rs143332484 C > T (R62H), and rs2234253 G > T (T96K). R47H was found only in the AD group, while T96K was present only in the controls. Although no significant association between TREM2 coding variants and LOAD susceptibility has been detected, the observed odds ratio (OR) of 3.69 for R47H carriers suggests an increased risk of LOAD for this variant in the Slovak population. Moreover, we also found a higher OR for the rs143332484-T allele in APOEε4 non-carriers (1.99) when compared to APOEε4 carriers (0.62). CONCLUSIONS: Our results suggest an impact of specific TREM2 rare coding variants on AD risk in the Slovak population.

Partial hydatidiform moles with unclear histopathological profile - importance of genetic diagnostics.

OBJECTIVE: The aim of the study was the genetic characterization of a set of cases with an unclear morphological profile of the placental tissue suspected of a partial hydatidiform mole. PATIENTS AND METHODS: This work presents the results of a genetic analysis of a group of 10 patients with various clinical manifestations of reproductive loss, where a partial hydatidiform mole was suspected on the basis of a histopathological examination. The composition of the genome of the products of conception was determined by short tandem repeats (STR) genotyping using a commercial kit;Devyser Compact v3 (Devyser). RESULTS AND CONCLUSIONS: Out of 10 analyzed cases, five had diandric monogynic triploid genome, characteristic for a partial mole. Aneuploidies of chromosomes 13, 18, 21, X and Y were excluded in four cases and Pataus syndrome was dia-gnosed in one case. In the case of an unclear histopathological profile, consultative DNA analysis (ideally STR genotyping) can significantly help the pathologist in the differential dia-gnosis of a partial mole. The histopathological profile of a partial hydatidiform mole may be in some cases incomplete and unclear, especially in the early weeks of gestation, which can lead to false negativity of the examination. On the other hand, other pathologies, for example aneuploides or digynic triploidy, may produce a histopathological profile similar to a partial mole, which leads to false positivity. Accurate dia-gnosis of a partial hydatidiform mole using molecular genetic methods contributes to the determination of adequate dispensary care for patients.

Transthyretin-related familial amyloid polyneuropathy (ATTR-FAP) in Poland - genetic and clinical presentation.

BACKGROUND: Transthyretin-related familial amyloid polyneuropathy (ATTR-FAP) is a rare, progressive, hereditary, highly disabling multisystem disorder. ATTR-FAP phenotypes differ according to the type of TTR mutation, geographic region and other as yet unidentified factors. The aim of this study was to establish the clinical and genetic characteristics of Polish patients. METHODS AND PATIENTS: Clinical data and necessary examinations were collected from patients diagnosed with ATTR-FAP at the Department of Neurology of Medical University of Warsaw between 1970 and 2019. RESULTS: 16 patients from eight unrelated families with five different TTR mutations were identified. The family with Val71Ala TTR mutation presented with early onset severe progressive polyneuropathy, with marked visual symptoms in a few patients. The next family with Ile73Val TTR mutation developed symptoms in middle age, and presented with mixed neuropathic and cardiologic phenotype. Four unrelated families were found to have the Phe33Leu TTR mutation with mixed neuropathic and cardiologic phenotype and late onset of symptoms. Other TTR mutations identified were: Val30Met and Asp38Val, both with late onset sensory, motor and autonomic neuropathy. CONCLUSION: Polish ATTR-FAP cases presented with heterogeneity typical for non-endemic areas. Phe33Leu TTR mutation was the most common, found in four unrelated families.

Neuropathology and biochemistry of early onset familial Alzheimer's disease caused by presenilin-1 missense mutation Thr116Asn.

The majority (~ 55%) of early onset familial Alzheimer disease (FAD) is caused by mutations in the presenilin 1 gene (PSEN1). Here, we describe a family with early onset FAD with a missense mutation in the PSEN1 gene (Thr116Asn). Five family members developed dementia in the third decade of life. One subject underwent autopsy. The onset of clinical symptoms was at the age of 37 years and the disease progressed rapidly. The clinical picture was characterised by progressive memory impairment, amnestic aphasia, and gait disturbances. Neuropathological assessment revealed widespread ß-amyloid (Thal phase 5) and tau (Braak stage 6) pathology. Abundant deposition of diffuse and cored plaques was distributed in cortical and subcortical areas, as well as in the cerebellum, while cotton wool plaques were observed mainly in the occipital cortex. Cerebral amyloid angiopathy was present throughout the brain. In the neocortex, tau pathology, especially neuropil threads, was more abundant in the frontal and occipital cortex and in the hippocampus. Proteomic analyses revealed that the pattern of sarkosyl-insoluble tau was similar to the one seen in sporadic AD. No α-synuclein or TDP-43 pathology was found either in cortical nor in subcortical areas. Here, we present the first comprehensive neuropathological and biochemical study of early onset FAD with a missense mutation Thr116Asn in the presenilin 1 gene. In contrast to other PS1-linked AD patients, the present subject developed cotton wool plaques which were not associated with spastic paraparesis.

Atypical Huntington's disease with the clinical presentation of behavioural variant of frontotemporal dementia.

Huntington's disease is an incurable, adult-onset, autosomal dominant inherited disorder caused by an expanded trinucleotide repeat (CAG). In this study, we describe a Huntington's disease patient displaying clinical symptoms of the behavioural variant of frontotemporal dementia in the absence of tremor and ataxia. The clinical onset was at the age of 36 years and the disease progressed slowly (18 years). Genetic testing revealed expanded trinucleotide CAG repeats in the Huntingtin gene, together with a Glu318Gly polymorphism in presenilin 1. Neuropathological assessment revealed extensive amyloid ß (Aß) aggregates in all cortical regions. No inclusions displaying hyperphosphorylated tau or phosphorylated transactive response DNA-binding protein 43 (TDP43) were found. A high number of p62 (sequestosome 1) immunopositive intranuclear inclusions were seen mainly in the cortex, while subcortical areas were affected to a lesser extent. Confocal microscopy revealed that the majority of p62 intranuclear lesions co-localised with the fused-in-sarcoma protein (FUS) immunostaining. The morphology of the inclusions resembled intranuclear aggregates in Huntington's disease. The presented proband suffered from Huntington's disease showed atypical distribution of FUS positive intranuclear aggregates in the cortical areas with concomitant Alzheimer's disease pathology.

A novel mutation in the PEX12 gene causing a peroxisomal biogenesis disorder.

The peroxisomal biogenesis disorders are autosomal recessive diseases morphologically characterised by lacking peroxisomes, biochemically by generalised deficiency of peroxisomal constituent and clinically manifested by serious health problems. Genes involved in the peroxisomal biogenesis are defined as the PEX genes encoding proteins called the peroxins. These peroxins are required for function in assembly of the peroxisomal membrane or in import of the enzymes into the peroxisomes. In this study we present a full overview of the clinical presentation, biochemical and molecular data of patient with Zellweger syndrome from Slovakia. We investigated biochemical metabolites using gas chromatography/mass spectrometry. The presence of causal ins/del mutations we identified by a Sanger sequencing and RFLP. We reported that the patient was a compound heterozygote for mutations in the gene PEX12: a 2-bp insertion (c.767_768dupAT) and a 2-bp deletion (c.887_888delTC). The first one mentioned is a novel mutation, which has not been reported before. Both mutations create a frameshift of the open reading frame which result a premature STOP codon and generate a complete loss of the C-terminal RING finger domain that is crucial for the correct import of proteins into peroxisomes. We found causal mutations responsible for a severe phenotype, and moreover we noted a novel mutation c.767_768dupAT that has not been reported before. The presence of mutations was studied in all family members, and the resulting data were successfully utilized for prenatal diagnosis.

Frequency of selected single nucleotide polymorphisms influencing the warfarin pharmacogenetics in Slovak population.

OBJECTIVE: Warfarin represents the most commonly prescribed oral anticoagulants, which functions as an antagonist of vitamin K, an essential factor of blood coagulation cascade. Warfarin has a narrow therapeutic index. An insufficient dose can cause failure of antithrombotic effect, and an overdose increases a risk of bleeding. It is known that variability in two genes (CYP2C9 and VKORC1) has a significant effect on individual response to warfarin dose. These polymorphisms influence more than one-third of known warfarin dose effect. Pharmacogenetics of warfarin is less affected by polymorphisms in the other genes such as CYP4F2, CYP2C19, and GGCX. MATERIAL AND METHODS: The frequency of selected single nucleotide polymorphisms including CYP2C9*2 (430C > T), CYP2C9*3 (1075A > C), VKORC1*2 (-1639G > A/1173C > T), VKORC1*3 (3730G > A), GGCX (12970C > G, 8016G > A), CYP2C19*2 (681G > A), and CYP4F2*3 (1297G > A) was tested in a control group consisting of 112 randomly selected individuals by allele-specific real-time PCR, restriction fragment length polymorphism, and bidirectional PCR allele-specific amplification. RESULTS AND DISCUSSION: The current results were statistically evaluated and compared with other populations. The presented results in Slovak population which is in Hardy-Weinberg equilibrium were compared with the prevalence in different countries. The incidence of selected polymorphisms in Slovak population correlates with Caucasians.

Differing clinical presentations of two unrelated cases of X-linked adrenoleukodystrophy with identical mutation Y296C in the ABCD1 gene.

OBJECTIVES: X-linked adrenoleukodystrophy is a genetically determined disorder that causes varying degrees of malfunction of the adrenal cortex and central nervous system. Our aim was to investigate the occurrence of known, or new, mutations in the ABCD1 gene in two unrelated patients with clinical suspicion of the adrenoleukodystrophy. METHODS: Two unrelated patients - the first with behavioral changes, the second with progressive cognitive deficit - underwent a clinical and genetic examination in order to establish a diagnosis and discover a possible mutation. RESULTS: In the first patient, a 47 year old man, the clinical examination showed dementia of the frontal type and spastic quadriparesis. The patient also suffered from adrenal insufficiency for 6 years. An MRI showed confluent hyperintensive lesions in FLAIR images in the frontal lobe of both hemispheres. The second patient, a 16 year old boy, suffered also from Addison's disease since the age of 9, and developed cognitive deficit in the course of one year. The MRI showed posterior atrophy and hyperintensive lesions in parietal and occipital lobes in T2WI. In both cases, genetic analyses showed a missense mutation at the codon 887 (A>G) in exon 1 of the ABCD1 gene, predicting the substitution Y296C in the ALD protein. CONCLUSION: We detected the same mutation of the ABCD1 gene in two unrelated patients with ALD. In the first case there was frontal lobe involvement, in the second case parieto-occipital involvement. Both pathologic involvement and clinical presentation differed in two cases of the same mutation.

Short tandem repeats genotyping of gestational choriocarcinoma - our experiences.

OBJECTIVE: This short communication demonstrates how short tandem repeat genotyping can identify the origin of gestational choriocarcinoma. MATERIALS AND METHODS: The origin of gestational choriocarcinoma in our three cases was determined using the short tandem repeats genotyping technique, which involved quantitative fluorescent PCR and fragmentation analysis. RESULTS: In Case 1 despite no medical history of molar pregnancy, DNA analysis indicated that the choriocarcinoma originated from a homozygous complete hydatidiform mole. We conclude, that the patient's complete abortion 10 years prior to the choriocarcinoma diagnosis was an undiagnosed complete hydatidiform mole. In Case 2 and Case 3 the clinically presumed origin of choriocarcinoma was confirmed. CONCLUSION: Determining the origin of choriocarcinoma is essential for clinical application, as it affects the FIGO scoring system for gestational trophoblastic neoplasia, which determines the patient's prognosis and treatment approach.

Dysfibrinogenemia and hypofibrinogenemia - Spectrum of pathogenic variants in Slovak patients.

INTRODUCTION: Congenital hypofibrinogenemia (CH) and congenital dysfibrinogenemia (CD) are rare coagulation disorders caused by quantitative or qualitative defects in the fibrinogen gene. The aim of this study was to characterize the genetic background and the clinical manifestations of congenital fibrinogen disorders in the patients from Slovakia registered at the National Haemophilia Centre. MATERIALS AND METHODS: Results of genetic analysis of the fibrinogen genes FGA, FGB and FGG using polymerase chain reaction followed by direct sequencing were evaluated in 36 patients. RESULTS: Molecular-genetic analysis revealed six novel variants - FGA c.923_968dup p.(Gly324Lysfs*44) and FGG c.1105C>T p.(His369Tyr) were identified in CD patients. In CH patients, in the FGG gene c.8G>A p.(Trp3*), c.823G>T p.(Glu275*) and c.323C>A p.(Ala108Asp) variants were detected. In the FGB gene c.1427C>T p.(Ser476Leu) was identified. CONCLUSION: This study is a positive contribution towards expanding knowledge about genetic variants in patients with congenital fibrinogen disorders.

Genetic and clinical characteristics including occurrence of testicular adrenal rest tumors in Slovak and Slovenian patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency.

Objective: To analyze the mutational spectrum, clinical characteristics, genotype-phenotype correlations, testicular adrenal rests tumor prevalence, and role of neonatal screening in congenital adrenal hyperplasia (CAH) patients from Slovakia and Slovenia. Design and methods: Data were obtained from 104 patients with CAH registered in Slovak and Slovenian databases. Low-resolution genotyping was performed to detect the most common point mutations. To detect deletions, conversions, point mutations, or other sequence changes in the CYP21A2 gene, high-resolution genotyping was performed. Genotypes were classified according to residual 21-hydroxylase activity (null, A, B, C). Results: 64% of the individuals had the salt-wasting form (SW-CAH), 15% the simple virilizing form (SV-CAH), and 21% the non-classic (NC-CAH). CYP21A2 gene deletion/conversion and c.293-13A/C>G pathogenic variant accounted together for 55.5% of the affected alleles. In SV-CAH p.Ile172Asn was the most common pathogenic variant (28.13%), while in NC-CAH p.Val282Leu (33.33%), CYP21A2 gene deletion/conversion (21.43%), c.293-13A/C>G (14.29%), Pro30Leu (11.90%). The frequency of alleles with multiple pathogenic variants was higher in Slovenian patients (15.83% of all alleles). Severe genotypes (0 and A) correlated well with the expected phenotype (SW in 94.74% and 97.3%), while less severe genotypes (B and C) correlated weaklier (SV in 50% and NC in 70.8%). The median age of SW-CAH patients at the time of diagnosis was 6 days in Slovakia vs. 28.5 days in Slovenia (p=0.01). Most of the Slovak patients in the cohort were detected by NBS. (24 out of 29). TARTs were identified in 7 out of 24 male patients, of whom all (100%) had SW-CAH and all had poor hormonal control. The median age at the diagnosis of TARTs was 13 years. Conclusion: The study confirmed the importance of neonatal screening, especially in the speed of diagnosis of severe forms of CAH. The prediction of the 21-OH deficiency phenotype was reasonably good in the case of severe pathogenic variants, but less reliable in the case of milder pathogenic variants, which is consistent compared to data from other populations. Screening for TARTs should be realized in all male patients with CAH, since there is possible remission when identified early.

Fabry Disease in Slovakia: How the Situation Has Changed over 20 Years of Treatment.

Fabry disease (FD, OMIM#301500) is a rare inborn error of the lysosomal enzyme α-galactosidase (α-Gal A, EC 3.2.1.22) and results in progressive substrate accumulation in tissues with a wide range of clinical presentations. Despite the X-linked inheritance, heterozygous females may also be affected. Hemizygous males are usually affected more severely, with an earlier manifestation of the symptoms. Rising awareness among health care professionals and more accessible diagnostics have positioned FD among the most-common inherited metabolic diseases in adults. An early and correct diagnosis of FD is crucial with a focus on personalised therapy. Preventing irreversible destruction of vital organs is the main goal of modern medicine. The aim of this study was to offer a complex report mapping the situation surrounding FD patients in Slovakia. A total of 48 patients (21 males, 27 females) with FD are registered in the Centre for Inborn Errors of Metabolism in Bratislava, Slovakia. In our cohort, we have identified three novel pathogenic variants in five patients. Three patients presented with the frameshift mutation c.736delA, and two others presented with the missense mutations c.203T>C, c.157A>C. Moreover, we present a new clinical picture of the pathogenic variant c.801+1G>A, which was previously described and associated with the renal phenotype.

[Hypophosphatasia--biochemical and clinical manifestations, molecular genetic principles]. / Hypofosfatázia--biochemické a klinické prejavy, molekulovo-genetická podstata.

Hypophosphatasia is a rare hereditary metabolic disorder accompanying deficit of tissue nonspecific serum alkaline phosphatase. The incidence of overt forms is estimated about 1:100000 live births. In the prenatal manifestation the disease may cause severe damage to the foetus with intrauterine death. In children there is a defect of mineralization with rickets signs and the subsequent hypercalcaemia a hypercalciuria may lead to death. In adults the main manifestation is osteomalacia, skeletal deformities and fractures, early arthritis. In severe forms the heredity is autosomal recessive type. In mild forms the heredity may be dominant or recessive. In two case reports we present clinical course of the disease in two adult sisters, where diagnosis of hypophosphatasia was first time confirmed in Slovak population using molecular genetic methods.

Allelic Distribution of Genes for Apolipoprotein E and MTHFR in Patients with Alzheimer's Disease and Their Epistatic Interaction.

BACKGROUND: Genetic risk factors play an important role in the pathogenesis of Alzheimer's disease (AD). However, the gene-gene interaction (epistasis) between specific allelic variants is only partially understood. OBJECTIVE: In our study, we examined the presence of the ɛ4 allele of apolipoprotein E (APOE) and the presence of C677T and A1298C (rs1801133 and rs1801131) polymorphisms in the methylenetetrahydrofolate reductase (MTHFR) gene in patients with AD and controls. We also evaluated the epistatic interaction between MTHFR and the APOE variants. METHODS: A total of 564 patients with AD and 534 cognitively unimpaired age-matched controls were involved in the study. RESULTS: The presence of the ɛ4 allele of APOE increases the risk of developing AD in a dose-dependent manner (OR 32.7: homozygotes, 15.6: homozygotes + heterozygotes, 14.3: heterozygotes). The combination of genotypes also increases the risk of developing AD in a dose-dependent manner: OR 18.3 (APOE 4/X and 4/4 + CT rs1801133), OR 19.4 (APOE 4/X and 4/4 + CT rs1801133 + AC rs1801131), OR 22.4 (APOE 4/X and 4/4 + TT rs1801133), and OR 21.2 (APOE 4/X and 4/4 + CC rs1801131). Homozygotes for variant alleles of MTHFR as well as patients with AD had significantly higher levels of homocysteine than homozygotes for standard alleles or controls. CONCLUSION: Homozygotes for APOE4 and carriers of APOE4 with TT genotype of rs1801133 were found to be at the highest risk of developing AD. These findings suggest that the epistatic interaction of specific gene variants can have a significant effect on the development of AD.

Smith-Lemli-Opitz syndrome - Fetal phenotypes with special reference to the syndrome-specific internal malformation pattern.

BACKGROUND: Autosomal-recessive SLOS is caused by mutations in the DHCR7 gene. It is defined as a highly variable complex of microcephaly with intellectual disability, characteristic facies, hypospadias, and polysyndactyly. Syndrome diagnosis is often missed at prenatal ultrasound and fetal autopsy METHODS: We performed autopsies and DHCR7 gene analyses in eight fetuses suspected of having SLOS and measured cholesterol values in long-term formalin-fixed tissues of an additional museum exhibit RESULTS: Five of the nine fetuses presented classical features of SLOS, including four cases with atrial/atrioventricular septal defects and renal anomalies, and one with additional bilateral renal agenesis and a Dandy-Walker cyst. These cases allowed for diagnosis at autopsy and subsequent SLOS diagnosis in two siblings. Two fetuses were mildly affected and two fetuses showed additional holoprosencephaly. These four cases and the exhibit had escaped diagnosis at autopsy. The case with bilateral renal agenesis presented a novel combination of a null allele and a putative C-terminus missense mutation in the DHCR7 gene CONCLUSIONS: In view of the discrepancy between the prevalence of SLOS among newborns and the carrier frequency of a heterozygous DHCR7 gene mutation, the syndrome-specific internal malformation pattern may be helpful not to miss SLOS diagnosis in fetuses at prenatal ultrasound and fetal autopsy.

Spectrum of DHCR7 mutations in Slovak patients with Smith-Lemli-Opitz syndrome and detection of common mutations by PCR-based assays.

The Smith-Lemli-Opitz syndrome (SLOS), an autosomal recessive disorder associated with multiple developmental malformations, is caused by a large spectrum of mutations in the DHCR7 gene. Mutations in the DHCR7 gene lead to a 7-dehydrocholesterol reductase deficiency, which is the final enzyme in the pathway of the cholesterol biosynthesis. Reduced cholesterol levels and elevated concentrations of its precursor 7-dehydrocholesterol in plasma and tissues are the major biochemical hallmarks of this disorder. In all patients a biochemical analysis of blood sterols using the gas chromatography/mass spectrometry was performed to confirm the clinical diagnosis of SLOS. We have also determined the mutational spectrum of DHCR7 gene in 17 Slovak patients. We identified six different mutations: nonsense mutation W151X and missense mutations V326L, L109P, G410S, R352Q, Y432C. Mutations W151X and V326L accounted for 76% of the SLOS alleles in Slovak population. The Slovak mutational spectrum is similar to that observed in other Central European countries. We also report simple polymerase chain reaction (PCR)-based assays that allow efficient and rapid mutation analysis.

Adult onset cerebral form of X-linked adrenoleukodystrophy with dementia of frontal lobe type with new L160P mutation in ABCD1 gene.

BACKGROUND: X-linked adrenoleukodystrophy is a genetically determined disorder that causes varying degrees of malfunction of the adrenal cortex and central nervous system and is characterized by abnormally high levels of very long chain fatty acid in tissues and body fluids. The gene ABCD1, responsible for X-ALD, has been mapped on chromosome Xq28. More than 500 different mutations have been reported but no correlation between genotype and phenotype has been found. OBJECTIVES: To investigate the occurrence of known or new mutations in the ABCD1 gene in patients with clinically and biochemical proven adrenoleukodystrophy. PATIENT AND METHODS: A 37-year-old patient with history of one-year progression of personality and behavioral changes such as, fluctuation of apathy and euphoria, perseveration, bizarre affect, and general disengagement, preliminarily assessed as adrenoleukodystrophy has undergone a clinical, biochemical and genetic examination in order to confirm the diagnosis and discover a possible mutation. RESULTS: The clinical examination has shown signs of the severe prefrontal syndrome, and a neurological examination disclosed deliberation signs and a spastic quadruparesis predominantly on the lower extremities. MRIs showed confluent hyperintensive lesions in T2 and FLAIR images in both hemispheres with severe progression over 6 to 12 months. Clinical findings referred to adrenoleukodystrophy, consecutively performed genetic analyses showed missense mutation at the codon 479 (T>C) in exon 1 of ABCD 1 gene, predicting the substitution L160P in ALD protein. The same mutation has also been found in patient's mother. CONCLUSION: We examined a patient with progressive development of early onset frontal lobe type dementia and upper motor neuron signs in which neuroimaging methods and biochemical tests refer to adrenoleukodystrophy. Genetic tests revealed a new mutation at position L160P in ALD protein.

Rapid and simple method for determination of Nepsilon-(carboxymethyl)lysine and Nepsilon-(carboxyethyl)lysine in urine using gas chromatography/mass spectrometry.

A new procedure was developed to determine in urine the concentrations of N(epsilon)-(carboxymethyl)lysine (CML) and N(epsilon)-(carboxyethyl)lysine (CEL), the major products of oxidative modification of glycated proteins, to assess levels of oxidative stress in physiological systems. The urine samples were acetonitrile-deproteinized, then derivatized by ethylchloroformate, and N(O,S)-ethoxycarbonyl ethyl esters of amino acids were analysed by isotope dilution gas chromatography/mass spectrometry. Recovery averaged 89%. Linearity was excellent (r = 0.998-0.999) in the 0.5-25 micromol/L range for CML and CEL. The limit of detection of this assay was 0.1 micromol/L (corresponding to 0.20 pmol of CML or CEL on column). Intra-day and inter-day precisions were likewise excellent, with relative standard deviations <4.63 and <6.15%, respectively. Accuracy of CML and CEL determination (15 micromol/L) was 2.9 and 5.9% of the estimated theoretical value. The time from obtaining the urine sample to determination of the concentration from the chromatographic peak was 80 min or less. This method is sensitive, reproducible, accurate, relatively cheap and very simple. It can be useful for laboratories involved in the diagnosis and monitoring of age-related chronic diseases.