Aberration of X chromosome in liver neoplasm detected by fluorescence in situ hybridization

BACKGROUND: A diverse range of cytogenetic alterations of autosomal chromosomes has been reported to date. However, few studies have addressed the abnormalities of X chromosome in hepatocellular carcinoma (HCC) except sporadic reports on the deletion of band F1 in X chromo- some , and the clonal analysis of methylation pattern of the X chromosome-linked human androgen receptor gene. Identification of specific X chromosome alterations during the course of neoplastic development would be essential to defining the genetic basis of HCC. Therefore, we studied the regularity of aberration of X chromosome in liver canc- er. METHODS: Hepatocarcinoma cellular lines and tumor tis- sues were detected respectively through DNA probes of X chromosome after fluorescence in situ hybridization ( FI- SH). RESULTS: Increased copies of X chromosome were ob- served in all samples, and four signals of hybridization were of the major type. CONCLUSIONS: Increased copy number of X chromo- some frequently occur in liver cancer. The relationship be- tween copy number of X chromosome and liver cancer genesis needs further investigation. This study is the first of its kind determining the copy number of X chromosome in liver cancer by using FISH.

Low XIST expression in Sertoli cells of Klinefelter syndrome patients causes high susceptibility of these cells to an extra X chromosome

Klinefelter syndrome(KS)is the most common genetic cause of human male infertility.However,the effect of the extra X chromosome on different testicular cell types remains poorly understood.Here,we profiled testicular single-cell transcriptomes from three KS patients and normal karyotype control individuals.Among the different somatic cells,Sertoli cells showed the greatest transcriptome changes in KS patients.Further analysis showed that X-inactive-specific transcript(XIST),a key factor that inactivates one X chromosome in female mammals,was widely expressed in each testicular somatic cell type but not in Sertoli cells.The loss of XIST in Sertoli cells leads to an increased level of X chromosome genes,and further disrupts their transcription pattern and cellular function.This phenomenon was not detected in other somatic cells such as Leydig cells and vascular endothelial cells.These results proposed a new mechanism to explain why testicular atrophy in KS patients is heterogeneous with loss of seminiferous tubules but interstitial hyperplasia.Our study provides a theoretical basis for subsequent research and related treatment of KS by identifying Sertoli cell-specific X chromosome inactivation failure.

Dynamic Spatial-temporal Expression Ratio of X Chromosome to Autosomes but Stable Dosage Compensation in Mammals

In the evolutionary model of dosage compensation,per-allele expression level of the X chromosome has been proposed to have twofold up-regulation to compensate its dose reduction in males(XY)compared to females(XX).However,the expression regulation of X-linked genes is still controversial,and comprehensive evaluations are still lacking.By integrating multi-omics datasets in mammals,we investigated the expression ratios including X to autosomes(X:AA ratio)and X to orthologs(X:XX ratio)at the transcriptome,translatome,and proteome levels.We revealed a dynamic spatial-temporal X:AA ratio during development in humans and mice.Meanwhile,by tracing the evolution of orthologous gene expression in chickens,platypuses,and opossums,we found a stable expression ratio of X-linked genes in humans to their autosomal orthologs in other species(X:XX1)across tissues and developmental stages,demonstrating stable dosage compensation in mammals.We also found that different epigenetic regulations contributed to the high tissue specificity and stage specificity of X-linked gene expression,thus affecting X:AA ratios.It could be concluded that the dynamics of X:AA ratios were attributed to the different gene contents and expression preferences of the X chromosome,rather than the stable dosage compensation.

Long noncoding RNA XIST:Mechanisms for X chromosome inactivation,roles in sex-biased diseases,and therapeutic opportunities

Sexual dimorphism has been reported in various human diseases including autoimmune diseases,neurological diseases,pulmonary arterial hypertension,and some types of cancers,although the underlying mechanisms remain poorly understood.The long noncoding RNA(lncRNA)X-inactive specific transcript(XIST)is involved in X chromosome inactivation(XCI)in female placental mammals,a process that ensures the balanced expression dosage of X-linked genes between sexes.XIST is abnormally expressed in many sex-biased diseases.In addition,escape from XIST-mediated XCI and skewed XCI also contribute to sex-biased diseases.Therefore,its expression or modification can be regarded as a biomarker for the diagnosis and prognosis of many sex-biased diseases.Genetic manipulation of XIST expression can inhibit the progression of some of these diseases in animal models,and therefore XIST has been proposed as a potential therapeutic target.In this manuscript,we summarize the current knowledge about the mechanisms for XIST-mediated XCI and the roles of XIST in sex-biased diseases,and discuss potential therapeutic strategies targeting XIST.

X chromosome inactivation and active X upregulation in therian mammals: facts, questions, and hypotheses

X chromosome inactivation is a mechanism that modulates the expression of X-linked genes in eutherian females(XX).Ohno proposed that to achieve a proper balance between X-linked and autosomal genes,those on the active X should also undergo a 2-fold upregula-tion.Although some support for Ohno's hypothesis has been provided through the years,recent genomic studies testing this hypoth-esis have brought contradictory results and fueled debate.Thus far,there are as many results in favor as against Ohno's hypothesis,depending on the nature of the datasets and the various assumptions and thresholds involved in the analyses.However,they have con-firmed the importance of dosage balance between X-linked and autosomal genes involved in stoichiometric relationships.These facts as well as questions and hypotheses are discussed below.

A Case of Two Full Sisters Share Identical Genotypes on the X Chromosome

X-chromosomal genetic markers are frequently employed in forensic parentage determination owing to their distinctive inheritance patterns.The kinship analysis revealed that two sisters who were not identical twins had identical genotypes on the X chromosome,encompassing 36 X-chromosomal short tandem repeats(X-STRs)and 29 X-chromosomal single-nucleotide polymorphisms(X-SNPs)that spanned the whole X chromosome from the p-telomere to the q-telomere.The identical X-STRs and X-SNPs in the daughters could be the result of linkage or a rare chance of occurrence.This highlights the need for careful analysis and interpretation when dealing with X chromosome markers and that in individual cases,even if two women share an allele at each locus,this does not necessarily mean that they are paternal sisters.The likelihood of random concordance due to maternal alleles must be taken into account.

An Uncommon Variant of Turner Syndrome in an African American Woman

The extra gonadal consequences of Turner’s Syndrome (TS) also pose risks to patients, namely cardiovascular. Clinicians should maintain a level of clinical suspicion for TS in patients with primary amenorrhea even without typical physical characteristics. Interestingly, TS has uncommon variant forms with varying degrees of clinical manifestations. Even so, all TS and TS variants maintain a high risk for cardiovascular events. Therefore, early TS diagnosis is of utmost importance. Here, we present a case of a young, African-American woman with primary amenorrhea with few overt clinical signs of TS. With high clinical suspicion, genetic testing is pursued and demonstrates the TS variant. This is important because variant forms have a similar increased risk of premature hypertension, diabetes, and aortic dissection.

Search for naive human pluripotent stem cells

Normal mouse pluripotent stem cells were originally derived from the inner cell mass(ICM) of blastocysts and shown to be the in vitro equivalent of those pre-implantation embryonic cells, and thus were called embryonic stem cells(ESCs). More than a decade later, pluripotent cells were isolated from the ICM of human blastocysts. Despite being called human ESCs, these cells differ significantly from mouse ESCs, including different morphology and mechanisms of control of pluripotency, suggesting distinct embryonic origins of ESCs from the two species. Subsequently, mouse pluripotent stem cells were established from the ICMderived epiblast of post-implantation embryos. These mouse epiblast stem cells(Epi SCs) are morphological and epigenetically more similar to human ESCs. This raised the question of whether cells from the human ICM are in a more advanced differentiation stage than their murine counterpart, or whether the available culture conditions were not adequate to maintain those human cells in their in vivo state, leading to a transition into Epi SC-like cells in vitro. More recently, novel culture conditions allowed the conversion of human ESCs into mouse ESC-like cells called nave(or ground state) human ESCs, and the derivation of nave human ESCs from blastocysts. Here we will review the characteristics of each type of pluripotent stem cells, how(and whether) these relate to different stages of embryonic development, and discuss the potential implications of nave human ESCs in research and therapy.

Study on the Relationship between Cytogenetics and Phenotypic Effect in Turner＇s Syndrome

The cytogenetics and clinical stigmata in 5 cases of Turner's syndrome were studied. Three of them were non-mosaic i(Xq) and two with partial monosome of a X chromosome short are (Xp21), whose DNA replication patterns of inactive X chromosome were analyzed by RBG technique. Results showed that differences between the replication patterns in cases of X chromosome deletion (Xp21) and normal females existed; that the behavior of abnormal X expressed nonrandom inactivation. It was suggested that the phenotype may be closely related with both X chromosome replication pattern and its inactivation behavior,which might be useful in genetic counselling.

Lack of evidence for leukocyte maternal microchimerism in primary biliary cirrhosis

AIM:It is reasonable to assume that microchimerism could also be involved in the induction of primary biliary cirrhosis (PBC).However,previous reports investigated only fetus-microchimerism in women patients.Maternal microchimerism has not been investigated until now. The current study aimed to clear either maternal microchimerism was involved in the pathogenesis of PBC or not. METHODS:We used fluorescence in situ hybridization on paraffin-embedded tissue (We called“Tissue-FiSH”.) to determine whether maternal cells infiltrated in male patients who were diagnosed as having PBC.Tissue-FiSH was performed by using both X and Y specific probes on the biopsy liver sample of 3 male PBC patients. RESULTS:Infiltrating lymphocytes demonstrated both X and Y signals in all 3 male patients. CONCLUSION:Maternal microchimerism dose not play a significant role in PBC.PBC may not relate to fetus and maternal microchimerism.

Effects of age on segregation of the X and Y chromosomes in cultured lymphocytes from Chinese men

Chromosome malsegregation in binucleated lymphocytes is a useful endpoint to evaluate age effect on genetic stability. However, the investigations on chromosome malsegregation in binucleated lymphocytes from Chinese are scarce. In this study, peripheral blood lym- phocytes were collected from 14 old （60-70 years） and 10 young （22-26 years） healthy Chinese men. To detect malsegregation of the sex chromosomes, multi-color fluorescence in situ hybridization （FISH） was performed on binucleated lymphocytes, cytokinesis-blocked by cytochalasin B at the first mitosis after phytohaemagglutinin stimulation. Compared with that in young men, a significant increase in frequencies of loss of chromosome X （9.2± 3.2‰ vs. 1.1 ± 0.9‰, P 〈 0.001） and Y （2.5 ± 1.9‰ vs. 0.2± 0.3‰, P 〈 0.001） was found in old men. Similarly, nondisjunction of chromosome X （16.5± 3.4‰ vs. 3.5 ± 1.1‰, P 〈 0.001） and Y （7.2 ± 2.6‰ vs. 2.4 ± 1.3‰, P 〈 0.001） occurred more frequently in old men than in young men. Regardless of donor＇s age, nondisjunction is more prevalent than loss for both chromosome X and Y. The frequencies of observed simultaneous malsegregation were relatively higher than the expected, suggest- ing an association between malsegregation. These results indicated that in Chinese men, malsegregation of the sex chromosomes increases with age in an associated fashion, and nondisjunction accounts for the majority of spontaneous chromosome malsegregation.

Two novel mutations of the retinitis pigmentosa GTPase regulator gene in two Chinese families with X-linked retinitis pigmentosa

Objective To detect mutations of the retinitis pigmentosa GTPase regulator (RPGR) gene in two Chinese X-linked retinitis pigmentosa families. Methods Fragments of exons 1-19 of the RPGR gene were amplified with intronic primers, using genomic DNA as template. The polymerase chain reaction (PCR) products were analysed by single-strand conformation polymorphism (SSCP) and direct sequencing. Mutations were identified by comparing DNA sequences of the patients with those of the normal controls.Results Two novel mutations, c1536delC and E332X, were identified in exons 12 and 9 of the RPGR gene in both families. Each mutation was the first mutation found in their respective exons. Both mutations were predicted to cause premature termination, which resulted in truncated proteins without normal functions of the RPGR products.Conclusions Both mutations are the genetic basis of the pathogenesis in the respective families. Our data might be helpful in analysing the function of the RPGR protein.

Genetic Polymorphisms of Nine X-STR Loci in Four Population Groups from Inner Mongolia, China

Nine short tandem repeat （STR） markers on the X chromosome （DXS101, DXS6789, DXS6799, DXS6804, DXST132, DXST133, DXS7423, DXS8378, and HPRTB） were analyzed in four population groups （Mongol, Ewenki, Oroqen, and Daur） from Inner Mongolia, China, in order to learn about the genetic diversity, forensic suitability, and possible genetic affinities of the populations. Frequency estimates, Hardy-Weinberg equilibrium, and other parameters of forensic interest were computed. The results revealed that the nine markers have a moderate degree of variability in the population groups. Most heterozygosity values for the nine loci range from 0.480 to 0.891, and there are evident differences of genetic variability among the populations. A UPGMA tree constructed on the basis of the generated data shows very low genetic distance betweent Mongol and Han （Xi＇an） populations. Our results based on genetic distance analysis are consistent with the results of earlier studies based on linguistics and the immigration history and origin of these populations. The minisatellite loci on the X chromosome studied here are not only useful in showing significant genetic variation between the populations, but also are suitable for human identity testing among Inner Mongolian populations.

Re-evaluation of the function of the male specific lethal complex in Drosophila

A set of proteins and noncoding RNAs, referred to as the male specific lethal （MSL） complex, is present on the male X chromosome in Drosophila and has been postulated to be responsible for dosage compensation of this chromosome -- the up-regulation of its expression to be equal to that of two X chromosomes in females. This hypothesis is evaluated in view of lesser known aspects of dosage compensation such as the fact that metafemales with three X chromosomes also have equal expression to normal females, which would require a down-regulation of each gene copy Moreover, when this complex is ectopically expressed in females or specifically targeted to a reporter in males, there is no increase in expression of the genes or targets with which it is associated. These observations are not consistent with the hypothesis that the MSL complex conditions dosage compensation. A synthesis is described that can account for these observations.

Genetic Polymorphism Investigation of 19 X‑STR Loci in the Han Population in Northern China

To investigate the genetic polymorphisms of 19 X‑STR loci in the Han population in Northern China,samples from 628 unrelated individuals(314 males and 314 females)were collected and 19 X‑STR loci were amplified by AGCU X19 STR System.A total of 270 different alleles were detected in 19 X‑STR loci.All loci were in Hardy−Weinberg equilibrium and there was only one pair of linkage loci(DXS10103‑DXS10101).There was no significant difference in allele frequency between male and female populations.The combined power of discrimination in males was 1–1.8667×10−13,while the combined power of discrimination in females was 1–3.6532×10−22.The combined mean paternity exclusion chance(CMEC)for X-chromosomal markers in father/daughter or mother/son duos Mean paternity exclusion chance(MECDesmarais Duo)was 1–5.1109×10−9.Moreover,the CMEC for X-chromosomal markers in trios involving daughters(MECDesmarais)was 1–2.0292×10−12.The compound amplification system composed of 19 X‑STR in this study showed high polymorphism in the Han population of Northern China,which had a high application value in difficult genetic relationship identification.

Identification of Half‑Sisters from Different Mothers by Autosomal and X Chromosomal Short Tandem Repeats:A Case Study

Complex kinship identification such as half‑sibling identification is a difficult task in forensic biology Here we represented an approach in dealing with half‑sisters from different mothers,with the combination of autosomal and X chromosomal short‑tandem repeats(STRs)data.X chromosomal STRs can offer additional information,especially in some cases where autosomal STRs alone may not provide enough information for an accurate opinion.In this case,half‑sister or unrelated relationship between two women(S_(1)and S_(2))with different mothers were distinguished.23 autosomal and 31 X chromosomal STRs of S_(1),S_(2),S_(1)’s mother(M1),S_(2)’s mother(M2)and S_(1)’s grandmother(G1)were profiled with three different commercial kits.As to X‑chromosome STRs,likelihood ratios(LRs)were calculated by FamLinkX with consideration of linkage,linkage disequilibrium,and mutations.When only the profiles of the two individuals(S_(1)and S_(2))were available,LRs between S_(1)and S_(2)were 1.1110×10^(2)based on 23 autosomal STRs and 3.2257 om107 based on 31 X chromosomal STRs.When the maternal genotypes were taken into consideration,LRs increased to 2.5297×10^(3)and 3.0563×10^(18).Therefore,both the DNA profiles of each mothers and X chromosomal STRs are important in dealing with the identification of half‑sisters from different mothers.

A Case of Maternal Half‑sisters Sharing Alleles at 18 X‑chromosomal Short Tandem Repeat Loci

Analysis of X‑chromosome short tandem repeats(STRs)is very helpful in deficiency paternity testing.Here,we reported a case of kinship analysis that showed a potentially erroneous inclusion of paternal sisters between two women.The two women shared alleles at 18 X‑chromosomal STR loci spanned from 14.76cM(DXS6807)to 184.19cM(DXS7423).When their relatives were not available for testing,biostatistical analysis for the 18 X‑chromosomal STR loci and 24 autosomal STR loci revealed the most possible relationship between the two women was paternal sisters.However,when the father of one woman was available,the other father‑daughter possibility was excluded.In the end,the likelihood ratio of STR marker and mitochondrial DNA(mtDNA)sequences confirmed the two women were maternal sisters.This case emphasizes a cautionary interpretation of X chromosomal marker in deficiency paternity cases with female offspring.Even though large parts of the X‑chromosome haplotypes shared by two females,additional relatives and extended DNA typing(such as mtDNA)may be needed further to ascertain whether they are paternal or maternal sisters.

Dual role of lipids for genome stability and pluripotency facilitates full potency of mouse embryonic stem cells

While Mek1/2 and Gsk3βinhibition("2i")supports the maintenance of murine embryonic stem cells(EsCs)in a homogenous naive state,prolonged culture in 2i results in aneuploidy and DNA hypomethylation that impairs developmental potential.Additionally,2i fails to support derivation and culture of fully potent female ESCs.Here we find that mouse ESCs cultured in 2i/LIF supplemented with lipid-rich albumin(AlbuMAx)undergo pluripotency transition yet maintain genomic stability and full potency over long-term culture.Mechanisticaily,lipids in AlbuMAx impact intracellular metabolism including nucleotide biosynthesis,lipid biogenesis,and TCA cycle intermediates,with enhanced expression of DNMT3s that prevent DNA hypomethylation.Lipids induce a formative-like pluripotent state through direct stimulation of Erk2 phosphorylation,which also alleviates X chromosome loss in female ESCs.Importantly,both male and female"all-ESc"mice can be generated from de novo derived ESCs using AlbuMAXbased media.Our findings underscore the importance of lipids to pluripotency and link nutrient cues to genome integrity in early development.

Role of Long Non-coding RNAs in Reprogramming to Induced Pluripotency

The generation of induced pluripotent stem cells through somatic cell reprogramming requires a global reorganization of cellular functions.This reorganization occurs in a multi-phased manner and involves a gradual revision of both the epigenome and transcriptome.Recent studies have shown that the large-scale transcriptional changes observed during reprogramming also apply to long noncoding RNAs(lncR NAs),a type of traditionally neglected RNA species that are increasingly viewed as critical regulators of cellular function.Deeper understanding of lncR NAs in reprogramming may not only help to improve this process but also have implications for studying cell plasticity in other contexts,such as development,aging,and cancer.In this review,we summarize the current progress made in profiling and analyzing the role of lncR NAs in various phases of somatic cell reprogramming,with emphasis on the re-establishment of the pluripotency gene network and X chromosome reactivation.