Use of multicolor fluorescence in situ hybridization to detect deletions in clinical tissue sections.

A variety of laboratory methods are available for the detection of deletions of tumor suppressor genes and losses of their proteins. The clinical utility of fluorescence in situ hybridization (FISH) for the identification of deletions of tumor suppressor genes has previously been limited by difficulties in the interpretation of FISH signal patterns. The first deletion FISH assays using formalin-fixed paraffin-embedded tissue sections had to deal with a significant background level of signal losses affecting nuclei that are truncated by the cutting process of slide preparation. Recently, more efficient probe designs, incorporating probes adjacent to the tumor suppressor gene of interest, have increased the accuracy of FISH deletion assays so that true chromosomal deletions can be readily distinguished from the false signal losses caused by sectioning artifacts. This mini-review discusses the importance of recurrent tumor suppressor gene deletions in human cancer and reviews the common FISH methods being used to detect the genomic losses encountered in clinical specimens. The use of new probe designs to recognize truncation artifacts is illustrated with a four-color PTEN FISH set optimized for prostate cancer tissue sections. Data are presented to show that when section thickness is reduced, the frequency of signal truncation losses is increased. We also provide some general guidelines that will help pathologists and cytogeneticists run routine deletion FISH assays and recognize sectioning artifacts. Finally, we summarize how recently developed sequence-based approaches are being used to identify recurrent deletions using small DNA samples from tumors.

Correction to: Use of multicolor fluorescence in situ hybridization to detect deletions in clinical tissue sections.

Figure 2 is incorrect in the original version of this article. The correct figure 2 is provided below.

Exercise Training-Induced Changes in MicroRNAs: Beneficial Regulatory Effects in Hypertension, Type 2 Diabetes, and Obesity.

MicroRNAs are small non-coding RNAs that regulate gene expression post-transcriptionally. They are involved in the regulation of physiological processes, such as adaptation to physical exercise, and also in disease settings, such as systemic arterial hypertension (SAH), type 2 diabetes mellitus (T2D), and obesity. In SAH, microRNAs play a significant role in the regulation of key signaling pathways that lead to the hyperactivation of the renin-angiotensin-aldosterone system, endothelial dysfunction, inflammation, proliferation, and phenotypic change in smooth muscle cells, and the hyperactivation of the sympathetic nervous system. MicroRNAs are also involved in the regulation of insulin signaling and blood glucose levels in T2D, and participate in lipid metabolism, adipogenesis, and adipocyte differentiation in obesity, with specific microRNA signatures involved in the pathogenesis of each disease. Many studies report the benefits promoted by exercise training in cardiovascular diseases by reducing blood pressure, glucose levels, and improving insulin signaling and lipid metabolism. The molecular mechanisms involved, however, remain poorly understood, especially regarding the participation of microRNAs in these processes. This review aimed to highlight microRNAs already known to be associated with SAH, T2D, and obesity, as well as their possible regulation by exercise training.

Complex Mosaic Ring Chromosome 11 Associated with Hemizygous Loss of 8.6 Mb of 11q24.2qter in Atypical Jacobsen Syndrome.

Jacobsen syndrome (JBS) is a contiguous gene deletion syndrome involving terminal chromosome 11q. The haploinsufficiency of multiple genes contributes to the overall clinical phenotype, which can include the variant Paris-Trousseau syndrome, a transient thrombocytopenia related to FLI1 hemizygous deletion. We investigated a boy with features of JBS using classic cytogenetic methods, FISH and high-resolution array CGH. The proband was found to have a mosaic ring chromosome 11 resulting in a hemizygous 11q terminal deletion of 8.6 Mb, leading to a copy number loss of 52 genes. The patient had a hemizygous deletion in the FLI1 gene region without apparent thrombocytopenia, and he developed diabetes mellitus type I, which has not previously been described in the spectrum of disorders associated with JBS. The relationship of some of the genes within the context of the phenotype caused by a partial deletion of 11q has provided insights concerning the developmental anomalies presented in this patient with atypical features of JBS.

Erratum to: Familial chromosomal translocation X; 22 associated with infertility and recurrent X mosaicism.

[This corrects the article DOI: 10.1186/s13039-016-0249-5.].

Familial chromosomal translocation X; 22 associated with infertility and recurrent X mosaicism.

BACKGROUND: Individuals with apparently balanced translocations, often, show no clinical findings. However, in meiosis, translocations tend to cause errors on chromosome disjunction and the ones involving sex chromosomes have particular implications for the phenotype. Male carriers of balanced X-autosome translocations are almost invariably infertile due to interruption of the spermatogenesis, but the mechanism is not fully understood. CASE PRESENTATION: In this case report, we performed a combination of classical cytogenetics (G-banding), molecular cytogenetics (fluorescence in situ hybridization and X-chromosome inactivation study), and cytogenomics (microarray-based comparative genomic hybridization) techniques for characterization of an inherited (X;22) translocation in a family originally referred for infertility investigation. Both proband and his sister are infertile and present the maternally inherited translocation. Interestingly, the maternal grandmother was mosaic for X chromosome monosomy suggesting that the t(X;22) in the proband's mother arose by errors at oogenesis. The presence of the same mosaicism of the X chromosome in the proband's aunt is consistent with this consideration. Array- CGH analysis showed no constitutional pathogenic gains or losses in the translocation carriers. The X-chromosome inactivation studies revealed that the translocated X;22 was active in 99.3% of cells in the mother and in 88% of cells in the daughter. We suggest that incomplete skewing of X inactivation (>97 %) of the daughter could justify the infertility. This study is the first description of recurrent mosaicism of the X chromosome associated with a familial X-autosome translocation. CONCLUSIONS: The phenotype of infertility was probably caused by disruption of spermatogenesis due to gametogenesis specific errors resulted from meiotic pairing and segregation anomalies on the translocated chromosomes.

Zika virus infection induces mitosis abnormalities and apoptotic cell death of human neural progenitor cells.

Zika virus (ZIKV) infection has been associated with severe complications both in the developing and adult nervous system. To investigate the deleterious effects of ZIKV infection, we used human neural progenitor cells (NPC), derived from induced pluripotent stem cells (iPSC). We found that NPC are highly susceptible to ZIKV and the infection results in cell death. ZIKV infection led to a marked reduction in cell proliferation, ultrastructural alterations and induction of autophagy. Induction of apoptosis of Sox2+ cells was demonstrated by activation of caspases 3/7, 8 and 9, and by ultrastructural and flow cytometry analyses. ZIKV-induced death of Sox2+ cells was prevented by incubation with the pan-caspase inhibitor, Z-VAD-FMK. By confocal microscopy analysis we found an increased number of cells with supernumerary centrosomes. Live imaging showed a significant increase in mitosis abnormalities, including multipolar spindle, chromosome laggards, micronuclei and death of progeny after cell division. FISH analysis for chromosomes 12 and 17 showed increased frequency of aneuploidy, such as monosomy, trisomy and polyploidy. Our study reinforces the link between ZIKV and abnormalities in the developing human brain, including microcephaly.

Smoking-induced chromosomal segregation anomalies identified by FISH analysis of sperm.

BACKGROUND: Numerical chromosome aberrations in gametes are directly related to infertility and aneuploid embryos. Previous studies have shown that toxic substances from cigarette smoke induce structural and numerical chromosomal aberrations in vitro and could potentially increase levels of aneusomy in sperm. Moreover, increased levels of aneusomy in sperm are correlated with low implantation rates, spontaneous abortions and fetal losses. Studies of chromosome 3 in sperm suggest it may be more prone to segregation anomalies than other autosomes, but there has been no systematic investigation of the incidence of disomy for chromosome 3 in sperm derived from donor male smokers. The objective of this study was to use FISH to evaluate the influence of smoking on the levels of disomy for chromosomes X and Y, and to determine whether disomy levels for chromosome 3 were elevated in sperm derived from male smokers. RESULTS: FISH analysis was used to evaluate the frequency of disomies of chromosomes 3, X, and Y in sperm of 10 smokers, compared to a control group of 7 non-smoking fertile men. All the subjects presented a normal somatic karyotype. There was a significant increase in the overall frequency of disomies in sperm derived from the smoking group (P< 0.0001). When each chromosome pair was analyzed individually, disomy of chromosome 3 in smokers was found to be more than twice that observed in the matched non-smoker control group. In addition we observed a higher frequencies of disomy of the X and Y chromosomes, indicating elevated levels of diploidy in the sperm from the smoking group. CONCLUSIONS: In this study we have shown that chromosome 3 may be susceptible to smoking-related segregation anomalies. Our results also suggest that errors can occur in both meiosis I and II, confirming the emerging literature that the male meiotic process may generally be affected by the genotoxic damage from tobacco use. Collectively, these findings provide additional evidence for enhancing tobacco control measures, and suggest that FISH analysis of chromosome 3 in sperm may be useful for monitoring smoking-induced segregation damage as part of the evaluation of infertile males.

Meiotic segregation and interchromosomal effect in the sperm of a double translocation carrier: a case report.

BACKGROUND: Infertility is a natural mechanism of selection intended to prevent the delivery of a child with malformations or mental retardation. Male infertility is closely related to chromosomal abnormalities. This study was focused on the analysis of meiotic segregation involving a Robertsonian translocation, 45,XY,der(13;13) [56]/45,XY,der(13;14) [44] and the evaluation of possible interchromosomal effects. RESULTS: Hybridisation with LSI 13q14 and subtelomere 14q probes and WCP13 SpectrumGreen and WCP14 SpectrumOrange probes showed a high proportion of unbalanced gametes, corresponding to 71.2% of the spermatozoa. The disomic frequencies of the sexual chromosomes and chromosome 18 of the patient were higher (5.28% and 2.55%, respectively) than those of the control (0.6% and 0.59%, respectively). CONCLUSION: Meiotic segregation studies in sperm are an important tool for genetic counselling of chromosomal aberrations, allowing for a prediction of the risks and consequent implications for the reproductive life. The patient with this rare translocation exhibited meiotic segregation fidelity, and a high rate of unbalanced gametes with disomic spermatozoa.