Ivana Joksic

Combined NGS approaches identify mutations in the intraflagellar transport gene IFT140 in skeletal ciliopathies with early progressive kidney Disease.

Ciliopathies are genetically heterogeneous disorders characterized by variable expressivity and overlaps between different disease entities. This is exemplified by the short rib‐polydactyly syndromes, Jeune, Sensenbrenner, and Mainzer‐Saldino chondrodysplasia syndromes. These three syndromes are frequently caused by mutations in intraflagellar transport (IFT) genes affecting the primary cilia, which play a crucial role in skeletal and chondral development. Here, we identified mutations in IFT140, an IFT complex A gene, in five Jeune asphyxiating thoracic dystrophy (JATD) and two Mainzer‐Saldino syndrome (MSS) families, by screening a cohort of 66 JATD/MSS patients using whole exome sequencing and targeted resequencing of a customized ciliopathy gene panel. We also found an enrichment of rare IFT140 alleles in JATD compared with nonciliopathy diseases, implying putative modifier effects for certain alleles. IFT140 patients presented with mild chest narrowing, but all had end‐stage renal failure under 13 years of age and retinal dystrophy when examined for ocular dysfunction. This is consistent with the severe cystic phenotype of Ift140 conditional knockout mice, and the higher level of Ift140 expression in kidney and retina compared with the skeleton at E15.5 in the mouse. IFT140 is therefore a major cause of cono‐renal syndromes (JATD and MSS). The present study strengthens the rationale for IFT140 screening in skeletal ciliopathy spectrum patients that have kidney disease and/or retinal dystrophy.

Dysfunctional telomeres in primary cells from Fanconi anemia FANCD2 patients

BackgroundFanconi anemia (FA) is characterized by sensitivity to DNA cross-linking agents, mild cellular, and marked clinical radio sensitivity. In this study we investigated telomeric abnormalities of non-immortalized primary cells (lymphocytes and fibroblasts) derived from FA patients of the FA-D2 complementation group, which provides a more accurate physiological assessment than is possible with transformed cells or animal models.ResultsWe analyzed telomere length, telomere dysfunction-induced foci (TIFs), sister chromatid exchanges (SCE), telomere sister chromatid exchanges (T-SCE), apoptosis and expression of shelterin components TRF1 and TRF2. FANCD2 lymphocytes exhibited multiple types of telomeric abnormalities, including premature telomere shortening, increase in telomeric recombination and aberrant telomeric structures ranging from fragile to long-string extended telomeres. The baseline incidence of SCE in FANCD2 lymphocytes was reduced when compared to control, but in response to diepoxybutane (DEB) the 2-fold higher rate of SCE was observed. In contrast, control lymphocytes showed decreased SCE incidence in response to DEB treatment. FANCD2 fibroblasts revealed a high percentage of TIFs, decreased expression of TRF1 and invariable expression of TRF2. The percentage of TIFs inversely correlated with telomere length, emphasizing that telomere shortening is the major reason for the loss of telomere capping function. Upon irradiation, a significant decrease of TIFs was observed at all recovery times. Surprisingly, a considerable percentage of TIF positive cells disappeared at the same time when incidence of γ-H2AX foci was maximal. Both FANCD2 leucocytes and fibroblasts appeared to die spontaneously at higher rate than control. This trend was more evident upon irradiation; the percentage of leucocytes underwent apoptosis was 2.59- fold higher than that in control, while fibroblasts exhibited a 2- h delay before entering apoptosis.ConclusionThe results of our study showed that primary cells originating from FA-D2 patients display shorten telomeres, elevated incidence of T-SCEs and high frequency of TIFs. Disappearance of TIFs in early response to irradiation represent distinctive feature of FANCD2 cells that should be examined further.

Improved diagnostic accuracy by using secondary ultrasound markers in the first-trimester screening for trisomies 21, 18 and 13 and Turner syndrome

The aim of this study was to examine the effectiveness of a combination of parameters at first‐trimester screening for fetal aneuploidies, including ultrasound assessment of the nasal bone (NB), blood flow in the ductus venosus (DV) and flow across the tricuspid valve.

Biological effects of Echinacea purpurea on human blood cells.

Biological Effects of Echinacea Purpurea on Human Blood Cells The aim of this study was to investigate radioprotective properties of Echinacea purpurea tablets in vivo. We analysed lymphocyte chromosome aberrations (CA), micronuclei (MN), apoptosis of leukocytes and haematological parameters in a group of radiation workers who were identified as carrying dicentric chromosomes in their lymphocytes. All radiation workers were taking two 275 mg Echinacea tablets b.i.d., according to a pharmacists recommendation. All parameters were analysed before and after the two-week treatment. At the end of the treatment lymphocyte CA frequency dropped significantly, and the number of apoptotic cells increased. The inverse lymphocyte-to-granulocyte ratio at the beginning of the study changed to normal at its end. In conclusion, biological effects observed after administration of Echinacea purpurea preparation suggest that it may be beneficial for the prevention of adverse health effects in workers exposed to ionising radiation. Biološki učinciI Echinacea purpurea na krvne stanice ljudi

A maternal deletion upstream of the imprint control region 2 in 11p15 causes loss of methylation and familial Beckwith-Wiedemann syndrome.

Beckwith–Wiedemann syndrome (BWS; OMIM #130650) is an overgrowth syndrome caused by different genetic or epigenetic alterations affecting imprinted regions on chromosome 11p15.5. Here we report a family with multiple offspring affected with BWS including giant omphalocoeles in which maternal transmission of a chromosomal rearrangement including an inversion and two deletions leads to hypomethylation of the imprint control region 2 (ICR2). As the deletion includes the promoter and 5′ part of the KCNQ1 gene, we suggest that transcription of this gene may be involved in establishing the maternal methylation imprint of the ICR2, which is located in intron 10 of KCNQ1.

BAC-Probes Applied for Characterization of Fragile Sites (FS)

Genomic instability tends to occur at specific genomic regions known as common fragile sites (FS). FS are evolutionarily conserved and generally involve late replicating regions with AT-rich sequences. The possible correlation between some FS and cancer-related breakpoints emphasizes on the importance of understanding the mechanisms of chromosomal instability at these sites. Although about 230 FS have already been mapped cytogenetically, only a few of them have been characterized on a molecular level. In this chapter, we provide a protocol for mapping of common FS using bacterial artificial chromosome (BAC) probes in fluorescence in situ hybridization (FISH) and suggest the usage of lymphocytes from Fanconi anemia patients as a model system. In the latter, rare FS are expressed much more frequently than in, for example, aphidicolin-induced blood lymphocyte preparations. Knowing the exact location of FS enables the molecular comparison of their location and breakpoints that appear during evolution, cancer development and inherited disorders.

First molecular-cytogenetic characterization of Fanconi anemia fragile sites in primary lymphocytes of FA-D2 patients in different stages of the disease

BackgroundFanconi anemia (FA) is a chromosomal instability syndrome characterized by increased frequency of chromosomal breakages, chromosomal radial figures and accelerated telomere shortening. In this work we performed detailed molecular-cytogenetic characterization of breakpoints in primary lymphocytes of FA-D2 patients in different stages of the disease using fluorescent in situ hybridization.ResultsWe found that chromosomal breakpoints co-localize on the molecular level with common fragile sites, whereas their distribution pattern depends on the severity of the disease. Telomere quantitative fluorescent in situ hybridization revealed that telomere fusions and radial figures, especially radials which involve telomere sequences are the consequence of critically shortened telomeres that increase with the disease progression and could be considered as a predictive parameter during the course of the disease. Sex chromosomes in FA cells are also involved in radial formation indicating that specific X chromosome regions share homology with autosomes and also could serve as repair templates in resolving DNA damage.ConclusionsFA-D2 chromosomal breakpoints co-localize with common fragile sites, but their distribution pattern depends on the disease stage. Telomere fusions and radials figures which involve telomere sequences are the consequence of shortened telomeres, increase with disease progression and could be of predictive value.