Ruth Mikelsaar

Cytogenetic analyses in infertile men.

In order to assess the significance of chromosome abnormalities and polymorphic chromosomal variants in male infertility, the results of cytogenetic studies of 90 patients (32 azoospermic and 58 oligozoospermic men) were compared with those of 30 control fertile men in Estonia. Total chromosome alterations were revealed in 47.8% of infertile men. Major chromosomal abnormalities had a 10-fold increase (13.4%) in infertile males (15.6% in azoospermics and 12% in oligozoospermics) compared to the control group. In azoospermics, the most prevalent were sex chromosomal abnormalities (47,XXY)(12%), whereas a Robertsonian translocation, t(13;14) was found in one patient (3.1). In contrast, in the oligospermia group the most frequent abnormalities were autosomal (10%), of which 6.9% were translocations. Sex chromosomal abnormalities were found very rarely (1.7%). Incidence of chromosomal variants was high (38%), but similar in all groups. The most frequently observed polymorphisms involved chromosome 9. In conclusion, chromosomal abnormalities found with a high frequency in infertile males are a major cause of male infertility, and justify the requirement of cytogenetic analysis in every infertile man.

Neonatal screening for congenital hypothyroidism in Estonia

Screening for congenital hypothyroidism was carried out by measuring thyroid stimulating hormone (TSH) on dried blood spots (mean + 2SD cut off value 12 μU/ml) by fluoroimmunoassay using DELFIA kits. A total of 20 021 infants were screened, and seven cases with congenital hypothyroidism were detected, giving an incidence of congenital hypothyroidism of 1:2860 (female:male ratio 6:1). In four of seven infants with congenital hypothyroidism (57%) the mother also had thyroid disease, supporting the importance of genetic factors as a cause of congenital hypothyroidism. Transient hyperthyrotropinaemia occurred in 654 infants (recall rate 3.3%). There was a significant association of transient hyperthyrotropinaemia only with cardiac failure at birth or caesarean section (p<0.01). Family studies showed no predisposition to thyroid diseases associated with a transient increase of TSH.

Epitope of titin A-band-specific monoclonal antibody Tit1 5 H1.1 is highly conserved in several Fn3 domains of the titin molecule. Centriole staining in human, mouse and zebrafish cells

BackgroundPreviously we have reported on the development of a new mouse anti-titin monoclonal antibody, named MAb Titl 5 H1.1, using the synthetic peptide N-AVNKYGIGEPLESDSVVAK-C which corresponds to an amino acid sequence in the A-region of the titin molecule as immunogen. In the human skeletal muscles, MAb Titl 5 H1.1 reacts specifically with titin in the A-band of the sarcomere and in different non-muscle cell types with nucleus and cytoplasm, including centrioles. In this report we have studied the evolutionary aspects of the binding of MAb Tit1 5 H1.1 with its target antigen (titin).ResultsWe have specified the epitope area of MAb Tit1 5 H1.1 by subpeptide mapping to the hexapeptide N-AVNKYG-C. According to protein databases this amino acid sequence is located in the COOH-terminus of several different Fn3 domains of the A-region of titin molecule in many organisms, such as human being, mouse, rabbit, zebrafish (Danio rerio), and even in sea squirt (Ciona intestinalis). Our immunohisto- and cytochemical studies with MAb Tit1 5 H1.1 in human, mouse and zebrafish tissues and cell cultures showed a striated staining pattern in muscle cells and also staining of centrioles, cytoplasm and nuclei in non-muscle cells.ConclusionsThe data confirm that titin can play, in addition to the known roles in striated muscle cells also an important role in non-muscle cells as a centriole associated protein. This phenomenon is highly conserved in the evolution and is related to Fn3 domains of the titin molecule. Using titin A-band-specific monoclonal antibody MAb Tit1 5 H1.1 it was possible to locate titin in the sarcomeres of skeletal muscle cells and in the centrioles, cytoplasm and nuclei of non-muscle cells in phylogenetically so distant organisms as Homo sapiens, Mus musculus and zebrafish (Danio rerio).

Two Novel Deletions (Array CGH Findings) in Pigment Dispersion Syndrome

Purpose: We report the first male with pigment dispersion syndrome and a balanced translocation t(10;15)(p11.1;q11.1). Methods: Cytogenetic analyses using Giemsa banding and FISH methods, and array CGH were performed. Results: Array CGH analyses did not show altered DNA sequences in the breakpoints of the translocation, but revealed two novel deletions in 2q22.1 and 18q22.1. Conclusion: We suppose that the coexistence of t(10;15) and pigment dispersion syndrome in our patient is a coincidence. The deletion in 2q22.1, where the gene LRP1B has been located, may play a major role in the dysembryogenesis of the eye and cause the disorder.

Balanced reciprocal translocation t(5;13)(q33;q12) and 9q31.1 microduplication in a man suffering from infertility and pollinosis

We describe the first case of two chromosomal abnormalities, balanced reciprocal translocation t(5;13)(q33;q12.1) and a microduplication in the region 9q31.1, in a man suffering from infertility and pollinosis. In the region 13q12.1 is located the TUBA3C (tubulin, alpha 3c) gene, which plays an important dynamic role in the motility of flagella. This case might support the opinion that haploinsufficiency of the TUBA3C gene could be the cause of sperm immotility and abnormal sperm morphology, resulting in infertility in the patient. Single-nucleotide polymorphism (SNP) array analysis revealed a novel 9q31.1 microduplication inherited from both parents, which contributes to the genomic instability.

Chromosome variant 1qh− and its influence on the 3D organization of chromosome 1 heterochromatin in interphase nucleus of patients with endometriosis

Chromosomes in the interphase nucleus are organized in chromosome territories with nonrandom chromosome positioning, which depend upon the chromosomal abnormality, gene density, size, transcriptional activity, early or late replication of sequences of DNA, and guanine–cytosine content (Monajembashi et al. 2005; Manvelyan et al. 2008; Klein et al. 2012). In spherical nuclei, such as lymphocytes, gene-dense chromosomes are found in the interior, genepoor chromosomes are found in the periphery of a nucleus (Monajembashi et al. 2005; Ferrai et al. 2010). The nuclear periphery is enriched for transcriptionally silent heterochromatin in most cell types, suggesting a mechanism for gene repression (Ferrai et al. 2010). But at present, it is known that these regions actively produce transcripts, which are vital to maintaining genome stability and proper cell division (Chen and Carmichael 2010; Hall et al. 2012). Disturbances in heterochromatin structure of chromosome 1 and its positioning in interphase nucleus have an important role in the regulation of gene expression (Cremer and Cremer 2001; Chen and Carmichael 2010), and have already been reported in two disorders, such as Roberts syndrome and immunodeficiency, centromeric region instability and facial anomalies (ICF) syndrome. Roberts syndrome, caused by the mutations of the ESCO2 gene, which encodes protein for sister chromatid cohesion during S phase, is characterized by the splitting of centromeric heterochromatin in metaphase cells. ICF is caused by the mutation in a DNA methyltransferase gene (DNMT3B), with hypomethylation

Small supernumerary marker chromosome (sSMC) derived from chromosome 22 in an infertile man with hypogonadotropic hypogonadism

We describe the first case of a supernumerary inv dup(22)(q11.1) in an infertile male with hypogonadotropic hypogonadism. This case supports the opinion that supernumerary inv dup(22)(q11.1) could play a role in male infertility. We suggest that the breakpoint in the region 22q11.1 and/or fourfold dosage of centromeric/pericentromeric sequences of the chromosome 22 may be the cause of hypogonadotropic hypogonadism resulting in impaired spermatogenesis and infertility in our patient.

Mosaic terminal del(19)(q13.33:) in a girl with seizures and mental retardation

Editor—Constitutional deletions of chromosome 19 are very rare.1 At present, only one case of 19p deletion2 and six cases of a de novo proximal deletion of 19q have been reported. The 19q deletions include an interstitial deletion del(19)(q12q13.1) and a submicroscopic de novo deletion of 19q13 resulting from t(X;19)(p21;q13) in a patient with Diamond-Blackfan anaemia and congenital anomalies.3 ,4 The analysis of familial and sporadic cases of Diamond-Blackfan anaemia showed four additional patients with de novo microdeletions that overlap the 19q13.2 region.5-7 No cases with a deletion of the terminal segment of 19q have been described so far. We report the first case of a constitutional terminal mosaic deletion of 19q with a breakpoint at band 19q13.33. The proband, a girl, was the result of the first pregnancy of healthy, unrelated, Estonian parents. At the childs birth, the mother was 20 and father 24 years old. The pregnancy was uneventful and the child was born at 42 weeks. Birth weight was 3920 g, length 51 cm, and head circumference (OFC) 35 cm. She was asphyxiated at birth (Apgar score was 2 at five minutes) and required artificial ventilation for two hours. On the third day she …

Titin A-band-specific monoclonal antibody Tit1 5H1.1. Cellular Titin as a centriolar protein in non-muscle cells.

We report the development of a new mouse anti-titin monoclonal antibody, named MAb Tit1 5H1.1, using the synthetic peptide corresponding to an amino acid sequence in the A-band of the titin molecule as immunogen. In the human skeletal muscle, MAb Tit1 5H1.1 reveals a clearly striated staining pattern, reacting with the A-band of the sarcomere. Electrophoretic, immunoblotting, and amino acid sequence analyses with ESI-MS/MS of human skeletal muscle tissue proved the target antigen of MAb Tit1 5H1.1 to be titin. The antibody reacts with titin also in non-muscle cells, producing a punctate pattern in cytoplasm and the nucleus. The most striking finding was a clear reaction of MAb Tit1 5H1.1 with centrioles in all cell types investigated so far. Immunocytochemical co-localization study with ninein-specific antibodies confirmed that the target antigen of MAb Tit1 5H1.1 is a centriole-associated protein. Experiments of the inhibition of synthesis of titin using titin siRNA duplex for the destruction of titin mRNA have shown a decreased staining of centrioles by MAb Tit1 5H1.1 in non-muscle cells and support the proposal that the target antigen of MAb is indeed titin. We suggest this anti-titin monoclonal antibody could be a valuable tool in the study of titin function and its subcellular location, both in muscle and non-muscle cells.

Alpha-1 antitrypsin phenotypes in patients with Klinefelter’s syndrome

Klinefelter’s syndrome (KS) is the most common human sex chromosome disorder, with a prevalence of 1 in 660 men. The phenotype is variable, but the most constant findings are small and firm testes, decreased testosterone level, infertility, eunuchoid body proportion, increased height, and learning disabilities, due to the presence of extra X chromosome(s) (Smyth and Bremner 1998). KS patients have frequently other diseases of lung, skin, liver and kidney (Morales et al. 1992; Swerdlow et al. 2005). Restrictive lung defects have been attributed to chest wall abnormalities, decreased respiratory muscle strength and increased chest wall compliance (Huseby and Petersen 1981). The likely cause is, decrease of lung compliance due to diminished elasticity of the lung matrix, but its biochemical cause is unknown (Morales et al. 1992). One of the causes could be the mutations in the alpha-1 antitrypsin (AAT) gene (SERPINE1), which is highly polymorphic, with more than 100 alleles identified so far. The alleles are categorized into normal, deficient and null variants on the basis of the plasma level and function of AAT. The protein phenotype is classified according to the ‘Pi’ (protease inhibitor) system, as defined by plasma isoelectric focusing. Normal Pi M types account for 95% of alleles in Caucasian individuals and are characterized by normal plasma levels. Pi X is a rare normal allele variant. Pi Z, S and null types are the most frequent AAT deficiency variants described in lung pathology, but Pi ZZ is associated with both pulmonary and liver diseases (Kaczor et al. 2007; Greene et al. 2008). AAT gene is located on chromosome 14q32.1 (Schroeder et al. 1985).