In this study, we aimed to explore the long noncoding RNA expression pattern in squamous cell lung cancer (SQCC) on a genome-wide scale. Total RNAs were extracted from 16 lung SQCC patients’ normal and matched lung cancer tissues by Trizol reagent. The expression level of genome-wide scale lncRNA and mRNA was determined by microarray. qRT-PCR was used to validate the lncRNA expression level in 47 patients. Data analyses were performed using R and Bioconductor. A total of 2,748 up and 852 down regulated probes were identified to be significantly and differentially expressed in tumor tissues. The annotation result of their co-expressed mRNAs showed that the most significantly related category of GO analysis was development and differentiation, while the most significantly related pathway was cell cycle. Subgroup analysis identified that 46 and 18 probes were specifically differentially expressed in smoking and moderately differentiated tumors, respectively. Our study indicated that clusters of lncRNAs were significantly and differentially expressed in SQCC compared with normal tissues in the same subject. They may exert a significant role in lung cancer development and could be potential targets for future treatment of SQCC.
Protein function has been observed to rely on select essential sites instead of requiring all sites to be indispensable. Small ubiquitin-related modifier (SUMO) conjugation or sumoylation, which is a highly dynamic reversible process and its outcomes are extremely diverse, ranging from changes in localization to altered activity and, in some cases, stability of the modified, has shown to be especially valuable in cellular biology. Motivated by the significance of SUMO conjugation in biological processes, we report here on the first exploratory assessment whether sumoylation related genetic variability impacts protein functions as well as the occurrence of diseases related to SUMO. Here, we defined the SUMOAMVR as sumoylation related amino acid variations that affect sumoylation sites or enzymes involved in the process of connectivity, and categorized four types of potential SUMOAMVRs. We detected that 17.13% of amino acid variations are potential SUMOAMVRs and 4.83% of disease mutations could lead to SUMOAMVR with our system. More interestingly, the statistical analysis demonstrates that the amino acid variations that directly create new potential lysine sumoylation sites are more likely to cause diseases. It can be anticipated that our method can provide more instructive guidance to identify the mechanisms of genetic diseases.
BACKGROUND: The nuclear factor-kB (NF-kB) family of transcriptional regulators are central mediators of the cellular inflammatory response. Although constitutive NF-kB signaling is present in most human tumours, mutations in pathway members are rare, complicating efforts to understand and block aberrant NF-kB activity in cancer. METHODS: To identify additional genetic alterations that drive ependymoma, we sequenced the whole genomes (WGS) of 41 tumours and matched normal blood, and the transcriptomes (RNAseq) of 77 tumours. The transforming significance of alterations were tested in mouse NSCs that we showed previously to be cells of origin of ependymoma. RESULTS: Here, we show that more than two thirds of supratentorial ependymomas contain oncogenic fusions between RELA, the principal effector of canonical NF-kB signalling, and an uncharacterized gene, C11orf95. In each case, C11orf95-RELA fusions resulted from chromothripsis involving chromosome 11q13.1. C11orf95-RELA fusion proteins translocated spontaneously to the nucleus to activate NF-kB target genes, and rapidly transformed neural stem cells—the cell of origin of ependymoma—to form these tumours in mice. CONCLUSIONS: Our data identify the first highly recurrent genetic alteration of RELA in human cancer, and the C11orf95-RELA fusion protein as a potential therapeutic target in supratentorial ependymoma. SECONDARY CATEGORY: Neuropathology & Tumor Biomarkers.
High HIV and syphilis incidence rates and prevalence among men who have sex with men in 2 cities of Jiangsu province indicate potential worsening of these epidemics in this population. Targeted intervention strategies are needed.
Background. Epidemics of human immunodeficiency virus (HIV) and syphilis among men who have sex with men (MSM) are major public health concerns in most parts of China. A dearth of information regarding the current trend of HIV in this hard-to-reach population in several regions including Jiangsu, coupled with the reemergence of syphilis, calls for an effort to understand the dynamics of the dual epidemic in this province.
Methods. To estimate the occurrence and burden of these 2 sexually transmitted diseases and the distribution of their potential sociobehavioral correlates among MSM in Jiangsu, 2 cohort studies were conducted in Yangzhou and Changzhou cities.
Results. Among 839 participants, 48% were married, 51.7% had sex with women in the last 6 months, and 25.5% did not use condoms during their last anal intercourse. The observed incidence of HIV was 13.59 and 12.62 and that of syphilis was 7.33 and 13.25 per 100 person-years among the participants of Yangzhou and Changzhou, respectively. The baseline prevalence of HIV and syphilis was 16.0% and 29.9% in Yangzhou and 13.6% and 14.9% in Changzhou, respectively.
Conclusions. Considerably high incidence and prevalence of HIV and syphilis among participants and their potential bridging role in transmitting these infections to the general population in Yangzhou and Changzhou cities calls for urgent effective intervention strategies.
HIV; STDs; MSM; China
The purpose of this study is to build a biological age (BA) equation combining telomere length with chronological age (CA) and associated aging biomarkers. In total, 139 healthy volunteers were recruited from a Chinese Han cohort in Beijing. A genetic index, renal function indices, cardiovascular function indices, brain function indices, and oxidative stress and inflammation indices (C-reactive protein [CRP]) were measured and analyzed. A BA equation was proposed based on selected parameters, with terminal telomere restriction fragment (TRF) and CA as the two principal components. The selected aging markers included mitral annulus peak E anterior wall (MVEA), intima-media thickness (IMT), cystatin C (CYSC), D-dimer (DD), and digital symbol test (DST). The BA equation was: BA = −2.281TRF + 26.321CYSC + 0.025DD − 104.419MVEA + 34.863IMT − 0.265DST + 0.305CA + 26.346. To conclude, telomere length and CA as double benchmarks may be a new method to build a BA.
Telomere length; Biological age; Chronological age; Aging markers
Linkage studies of alcoholism have implicated several chromosome regions, leading to the successful identification of susceptibility genes, including ADH4 and GABRA2 on chromosome 4. Quantitative endophenotypes that are potentially closer to gene action than clinical endpoints offer a means of obtaining more refined linkage signals of genes that predispose alcohol use disorders (AUD). In this study we examine a self-reported measure of the maximum number of drinks consumed in a 24-hour period (abbreviated Max Drinks), a significantly heritable phenotype (h2 = 0.32 ± 0.05; P = 4.61 × 10−14) with a strong genetic correlation with AUD (ρg = 0.99 ± 0.13) for the San Antonio Family Study (n = 1,203). Genome-wide SNPs were analyzed using variance components linkage methods in the program SOLAR, revealing a novel, genome-wide significant QTL (LOD = 4.17; P = 5.85 × 10−6) for Max Drinks at chromosome 6p22.3, a region with a number of compelling candidate genes implicated in neuronal function and psychiatric illness. Joint analysis of Max Drinks and AUD status shows that the QTL has a significant non-zero effect on diagnosis (P = 4.04 × 10−3), accounting for 8.6% of the total variation. Significant SNP associations for Max Drinks were also identified at the linkage region, including one, rs7761213 (P = 2.14 × 10−4), obtained for an independent sample of Chinese families. Thus, our study identifies a potential risk locus for AUD at 6p22.3, with significant pleiotropic effects on the heaviness of alcohol consumption that may not be population specific.
AUD; alcohol dependence; variance components linkage analysis; pleiotropy; endophenotype ranking value (ERV); SNP association
AIM: To comprehensively understand the underlying molecular events accounting for aberrant Wnt signaling activation in hepatocellular carcinoma (HCC).
METHODS: This study was retrospective. The HCC tissue specimens used in this research were obtained from patients who underwent liver surgery. The Catalogue of Somatic Mutations in Cancer (COSMIC) database was searched for the mutation statuses of CTNNB1, TP53, and protein degradation regulator genes of CTNNB1. Dual-luciferase reporter assay was performed with TOP/FOP reporters to detect whether TP53 gain-of-function (GOF) mutations could enhance the transcriptional activity of Wnt signaling. Methylation sensitive restriction enzyme-quantitative PCR was used to explore the methylation status of CpG islands located in the promoters of APC, SFRP1, and SFRP5 in HCCs with different risk factors. Finally, nested-reverse transcription PCR was performed to examine the integration of HBx in front of LINE1 element and the existence of HBx-LINE1 chimeric transcript in Hepatitis B virus-related HCC. All results in this article were analyzed with the software SPSS version 19.0 for Windows, and different groups were compared by χ2 test as appropriate.
RESULTS: Based on the data from COSMIC database, compared with other solid tumors, mutation frequency of CTNNB1 was significantly higher in HCC (P < 0.01). The rate of CTNNB1 mutation was significantly less frequent in Hepatitis B virus-related HCC than in other etiologies (P < 0.01). Dual-luciferase reporter system and TOP/FOP reporter assays confirmed that TP53 GOF mutants were able to enhance the transcriptional ability of Wnt signaling. An exclusive relationship between the status of TP53 and CTNNB1 mutations was observed. However, according to the COSMIC database, TP53 GOF mutation is rare in HCC, which indicates that TP53 GOF mutation is not a reason for the aberrant activation of Wnt signaling in HCC. APC and AXIN1 were mutated in HCC. By using methylation sensitive restriction enzyme-quantitative PCR, hypermethylation of APC was detected in HCC with different risk factors, whereas SFRP1 and SFRP5 were not hypermethylated in any of the HCC etiologies, which indicates that the mutation of APC and AXIN1, together with the methylation of APC could take part in the overactivation of Wnt signaling. Nested-reverse transcription PCR failed to detect the integration of HBx before the LINE1 element, or the existence of an HBx-LINE1 chimeric transcript, suggesting that integration could not play a role in the aberrant activation of Wnt signaling in HCC.
CONCLUSION: In HCC, genetic/epigenetic aberration of CTNNB1 and its protein degradation regulators are the major cause of Wnt signaling overactivation.
β-catenin; CTNNB1; Hepatitis B virus; Hepatocellular carcinoma; TP53; Wnt signaling
Near-field optical techniques exploit light-matter interactions at small length scales for mechanical sensing and actuation of nanomechanical structures. Here, we study the optical interaction between two mechanical oscillators—a plasmonic nanofocusing probe-tip supported by a low frequency cantilever, and a high frequency nanomechanical resonator—and leverage their interaction for local detection of mechanical vibrations. The plasmonic nanofocusing probe provides a confined optical source to enhance the interaction between the two oscillators. Dynamic perturbation of the optical cavity between the probe-tip and the resonator leads to nonlinear modulation of the scattered light intensity at the sum and difference of their frequencies. This double-frequency demodulation scheme is explored to suppress unwanted background and to detect mechanical vibrations with a minimum detectable displacement sensitivity of 0.45 pm/Hz1/2, which is limited by shot noise and electrical noise. We explore the demodulation scheme for imaging the bending vibration mode shape of the resonator with a lateral spatial resolution of 20 nm. We also demonstrate the time-resolved aspect of the local optical interaction by recording the ring-down vibrations of the resonator at frequencies of up to 129 MHz. The near-field optical technique is promising for studying dynamic mechanical processes in individual nanostructures.
Ewing sarcoma is a primary bone tumor initiated by EWSR1–ETS gene fusions. To identify secondary genetic lesions that contribute to tumor progression, we performed whole-genome sequencing of 112 Ewing sarcoma samples and matched germline DNA. Overall, Ewing sarcoma tumors had relatively few single-nucleotide variants, indels, structural variants and copy-number alterations. Apart from whole chromosome arm copy-number changes, the most common somatic mutations were detected in STAG2 (17%), CDKN2A (12%), TP53 (7%), EZH2, BCOR, and ZMYM3 (2.7% each). Strikingly, STAG2 mutations and CDKN2A deletions were mutually exclusive, as confirmed in Ewing sarcoma cell lines. In an expanded cohort of 299 patients with clinical data, we discovered that STAG2 and TP53 mutations are often concurrent and are associated with poor outcome. Finally, we detected subclonal STAG2 mutations in diagnostic tumors and expansion of STAG2 immuno-negative cells in relapsed tumors as compared with matched diagnostic samples.
Ewing sarcoma; genomics; mutations; whole genome sequencing; prognostic
Metastasis is a leading cause of mortality for osteosarcoma patients. The molecular pathological mechanism remains to be elucidated. In the previously study, we established two osteosarcoma cell lines with different metastatic potentials. Differential expressed genes and proteins regarding metastatic ability have been identified. MicroRNAs are important regulators in tumorigenesis and tumor progression. In this study, microRNA microarray was used to assess the differential expressed miRNAs level between these two cell lines. One of the top ranked miRNAs-miR-195 was identified highly expressing in lowly metastatic cells. It was showed that over-expression of miR-195 substantially inhibits migration and invasion of osteosarcoma cells in vitro and pulmonary metastasis formation in vivo. Meanwhile, CCND1 was identified as the target gene of miR-195 and further studied. More importantly, Using real-time PCR, we evaluated the expression of miR-195 and CCND1 in osteosarcoma samples from 107 frozen biopsy tissues and 99 formalin- or paraformalin-fixed, paraffin-embedded (FFPE) tissues. Results indicated lowly expressed miR-195 or highly CCND1 correlated with positive overall survival and their expression inverse relate to each other. In summary, our study suggests miR-195 function as a tumor metastasis suppressor gene by down-regulating CCND1 and can be used as a potential target in the treatment of osteosarcoma.
microRNA; miR-195; osteosarcoma; CCND1; metastatic
Traditional Chinese medicine (TCM) is an important treatment for male infertility, and its application to therapy is dependent on differentiation of TCM syndromes. This study aims to investigate the changes in metabolites and metabolic pathways in infertile males with Kidney-Yang Deficiency syndrome (KYDS) via metabolomics approaches. Seminal plasma samples were collected from 18 infertile males with KYDS and 18 fertile males. Liquid chromatography and mass spectrometry were used to characterize metabolomics profiles. Principal component analysis (PCA), partial least squares-discriminate analysis (PLS-DA), and pathway analysis were used for pattern recognition and metabolite identification. PCA and PLS-DA results differentiated the two groups of patients. Forty-one discriminating metabolites (18 in positive mode and 23 in negative mode) were identified. Seven metabolites were related to five potential metabolic pathways associated with biosynthesis and metabolism of aromatic amino acids, tricarboxylic acid cycle, and sphingolipid metabolism. The changes in metabolic pathways may play an important role in the origin of KYDS-associated male infertility. Metabolomics analysis of seminal plasma may be used to differentiate TCM syndromes of infertile males, but further research must be conducted.
Diabetic patients with acute coronary syndrome (ACS) are at higher risk of poor outcome than are non-diabetic patients with ACS. Few studies have focused on sex-related ACS incidence, ACS-related mortality or risk factors to affects sex specific ACS in Chinese with Type 2 diabetes mellitus (T2DM). Based on a hospital-based cohort of Chinese patients with T2DM, we aimed to investigate whether there was sex difference in ACS or ACS-related mortality or risk factors of ACS.
Totally 2,135 Hong Kong Chinese with T2DM were recruited during 1994-1996 and followed up until August 2012. We systematically analyzed sex-related ACS incidence and ACS-related mortality and risk factors with χ2-squared test, descriptive statistics and survival analysis.
Regular follow-up was completed in 2,105 subjects (98.6%), with a median period of 14.53 years. The occurrence of ACS was recorded among 414 patients (19.7%) and ACS-related death among 104 patients (4.9%). ACS incidences increased with age in both men and women, and men had a higher prevalence of ACS than women across different age categories and different follow-up periods (log rank χ2=20.32, P<0.001). The transition of ACS incidences from slow to rapid increase were about 5 years earlier in men (at 51-55 years) than in women (55-60 years). Among ACS patients, cumulative ACS-related mortalities was similar between men and women (log rank χ2=0.063, P=0.802). Besides age and albuminuria, different profiles of risk factors accounted for the occurrence of ACS between men and women.
Our findings demonstrated sex differences in ACS incidence and risk factors, but not in ACS-related mortality in Chinese patients withT2DM. These findings suggest that screening and prevention campaigns should be optimized for men and women, which may help to identify diabetic patients at higher risk of coronary heart disease.
Toll-like receptor 7 (TLR7) is activated by single-stranded RNA and synthetic imidazoquinoline components, and induces interferon production. In this study, we cloned the TLR7 gene from King pigeon (Columba livia). The TLR7 open reading frame is 3144 bp and encodes a 1047-amino acid protein, consisting of a canonical TLR composition with 15 leucine-rich repeats (LRRs). Amino acid-inserting modifications were found at position 15 of LRR2, LRR11, LRR13, and LRR14 and position 10 of LRR10. The tissue distribution of pigeon TLR7 suggests that immune-associated tissues, especially the spleen and liver, have high TLR7 expression. HEK293T cells transfected with pigeon TLR7 plasmid responded to the agonist R848, indicating a functional TLR7 homolog. Following R848 stimulation of pigeon peripheral blood mononuclear cells, the levels of IFN-γ, IL-6, IL-8, CCL5, and IL-10 mRNA, assessed using quantitative real-time PCR, were significantly up-regulated. After Newcastle disease virus vaccine strain LaSota inoculation and agonist R848 injection, the level of TLR7 mRNA in the spleen of pigeons increased significantly in the R848-injected group, but decreased in the LaSota-inoculated group at three day post-infection (d.p.i.). The mRNA levels of inflammatory cytokines and chemokines were significantly upregulated in both LaSota-inoculated and R848-injected groups. Triggering pigeon TLR7 leads to robust up-regulation of inflammatory cytokines and chemokines, suggesting an important role in the innate immune response.
TLR7; pigeon; identification; characterization; immune function
The mechanism by which overexpression of hexokinase 2 (HK2) indicates locally advanced cervical squamous cell carcinoma (LACSCC) with radio-resistance is still unknown despite being an independent biomarker of poor prognosis. Here, we retrospectively analyzed 132 female patients receiving radiotherapy for cervical squamous cell carcinoma including 85 radiation-sensitive cases and 47 radiation-resistant cases. The expression of HK2 was examined by immunohistochemistry. The percentage of high HK2 expression in the radiation-resistant group differed from the radiation-sensitive group with statistical significance (P < 0.001) even if divided into three subgroups including a lower 5-year progression free survival group (PFS) for comparison (P < 0.001). The Kaplan Meier curve analysis showed that there were differences between the two groups (P < 0.001). Therefore, this study proves a close relationship between HK2 expression and radio-resistance. Multivariate Cox regression analysis implied that HK2 was an independent prognostic indicator of cervical squamous carcinoma (HR (95% CI), 2.940 (1.609, 1.609); P = 0.002).
Locally advanced cervical squamous cell carcinoma; radiation resistance; glycolysis; HK2; immunohistochemistry
Purpose. To subclassify parapharyngeal extension in nasopharyngeal carcinoma (NPC) and investigate its prognostic value and staging categories based on magnetic resonance imaging (MRI). Methods and Materials. Data from 1504 consecutive NPC patients treated with definitive-intent radiotherapy were analyzed retrospectively. Sites of parapharyngeal extension were defined by MRI. Overall survival (OS), local relapse-free survival (LRFS), and distant metastasis-free survival (DMFS) were calculated by the Kaplan-Meier method and compared with the log-rank test. Hazard consistency and hazard discrimination were determined by multivariate analysis with Cox proportional hazards models. Results. 1104 patients (73.4%) had parapharyngeal extension; 1.7–63.8% had involvement of various anatomic sites. The hazard ratio for death was significantly higher with extensive parapharyngeal extension (lateral pterygoid muscle of masticator space and beyond or parotid space) than with mild extension (medial pterygoid muscle of masticator space, or carotid, prestyloid, and prevertebral or retropharyngeal space). OS, LRFS, and DMFS with extensive parapharyngeal extension were similar to those in T4 disease; OS, LRFS, and DMFS with mild parapharyngeal extension were significantly higher than in those T3 disease (all P ≤ 0.015). Conclusions. Parapharyngeal extension in NPC should be subclassified as mild or extensive, which should be regarded as stages T2 and T4 diseases, respectively.
Sustained, high level transgene expression in mammalian cells, especially stem cells, may be desired in many cases for studying gene functions. Traditionally, stable transgene expression has been accomplished by using retroviral or lentiviral vectors. However, such viral vector-mediated transgene expression is often at low levels and can be reduced over time due to low copy numbers and/or chromatin remodeling repression. The piggyBac transposon has emerged as a promising non-viral vector system for efficient gene transfer into mammalian cells. Despite its inherent advantages over lentiviral and retroviral systems, piggyBac system has not been widely used, at least in part due to the limited availability of piggyBac vectors with manipulation flexibilities. Here, we seek to optimize piggyBac-mediated transgene expression and generate a more efficient, user-friendly piggyBac system. By engineering a panel of versatile piggyBac vectors and constructing recombinant adenoviruses expressing piggyBac transposase (PBase), we demonstrate that adenovirus-mediated PBase expression significantly enhances the integration efficiency and expression level of transgenes in mesenchymal stem cells and osteosarcoma cells, compared to that obtained from co-transfection of the CMV-PBase plasmid. We further determine the drug selection timeline to achieve optimal stable transgene expression. Moreover, we demonstrate that the transgene copy number of piggyBac-mediated integration is approximately 10 times higher than that mediated by retroviral vectors. Using the engineered tandem expression vector, we show that three transgenes can be simultaneously expressed in a single vector with high efficiency. Thus, these results strongly suggest that the optimized piggyBac system is a valuable tool for making stable cell lines with sustained, high transgene expression.
piggyBac transposon; piggyBac transposase; stable transgene expression; mesenchymal stem cells; retroviral vectors; transposition
In China, the HIV/AIDS epidemic is expanding among men who have sex with men (MSM). As independent risk factors of HIV infection, the epidemics of Chlamydia (CT) and Gonorrhea (NG) in MSM were not well studied, particular for the risk factors of these infectious. The objectives of current reported study were to understand the dynamics of HIV and other sexual transmitted infections (STIs) among MSM in Jiangsu, China, and to measure factors that correlated with STIs.
In order to gain more participants, a multisite cross-sectional study design was used in our study, by using convenience-sampling to recruit MSM in two Changzhou and Yangzhou, Jiangsu, China, between the July and October of 2009.
In this comprehensive survey involving MSM in two cities of Jiangsu province of China, the prevalence of STIs of CT (6.54%), NG (3.63%), syphilis (20.34%) and HIV (11.62%) were measured. Overall, the STIs prevalence (CT, NG or syphilis) for the participants in our study was 26.39%, meanwhile, 3.4% (14 out of the 413) participants had at least two kinds of STIs. Meeting casual partners at parks, public restrooms or other public areas, having had anal sex with men in the past six months, having had STI symptoms in the past year were positively correlated with STIs positive, with adjusted ORs of 4.61(95%CI 1.03–20.75), 1.91(95%CI 1.14–3.21) and 2.36(95%CI 1.07,5.24).
Our study findings reiterate the fact that Chinese MSM are highly susceptible to acquiring syphilis, CT, NG and HIV, and there is an urgent need for intervention targeted towards this population. Behavioral measures should constitute an important part of the targeted intervention. Furthermore, the already implemented preventive and diagnostic services for HIV should be expanded to include syphilis CT and NG, too.
A great number of naturally occurring oligosaccharides and oligosaccharide esters have been isolated from traditional Chinese medicinal plants, which are used widely in Asia and show prominent curative effects in the prevention and treatment of kinds of diseases. Numerous in vitro and in vivo experiments have revealed that oligosaccharides and their esters exhibited various activities, including antioxidant, antidepressant, cytotoxic, antineoplastic, anti-inflammatory, neuroprotective, cerebral protective, antidiabetic, plant growth-regulatory, and immunopotentiating activities. This review summarizes the investigations on the distribution, chemical structures, and bioactivities of natural oligosaccharides and their esters from traditional Chinese medicines between 2003 and 2013.
Activation of microglia plays a crucial role in immune and inflammatory processes after ischemic stroke. Microglia is reported with two opposing activated phenotypes, namely, classic phenotype (M1) and the alternative phenotype (M2). Inhibiting M1 while stimulating M2 has been suggested as a potential therapeutic approach in the treatment of stroke.
In this study, we indicated that a novel natural anti-oxidant extracted from the Chinese plant Hopea hainanensis, malibatol A (MA), decreased the infarct size and alleviated the brain injury after mice middle cerebral artery occlusion (MCAO). MA inhibited expression inflammatory cytokines in not only MCAO mice but also lipopolysaccharide (LPS)-stimulated microglia. Moreover, treatment of MA decreased M1 markers (CD16, CD32, and CD86) and increased M2 markers (CD206, YM-1) while promoting the activation of nuclear receptor PPARγ.
MA has anti-inflammatory effects in MCAO mice in a PPARγ-dependent manner, making it a potential candidate for stroke treatment.
Experimental stroke; Inflammation; Malibatol A; Microglia; M1/M2; PPARγ
MicroRNAs play an important role in the development and progression of Ewing's sarcoma (ES). Especially, the expression of let-7a has been reported to be significantly downregulated in various cancers, and can affect the initiation and maintenance of tumor progression. However, the relative effects of let-7a on ES cells and relative mechanisms are largely unknown. In this study, we identified the underexpression of let-7a in human ES cells comparing with the human mesenchymal stem cells. Then, we sought to compensate for its loss through exogenous transfection with let-7a mimic into ES cell lines A673 and SK-ES-1. Restored let-7a expression inhibited cell proliferation, migration, as well as invasion; arrested cell cycle progression; and induced cell apoptosis of both cell lines. Moreover, bioinformatic prediction suggested that cyclin-dependent kinase 6 (CDK6), which is overexpressed and functions as an oncoprotein in ES cells, is a putative target gene of let-7a. Using mRNA and protein expression analysis and luciferase assays, we further identified the target role of CDK6. Finally, we found that restored CDK6 expression in ES cells that had been treated with let-7a mimic before could partly dampen let-7a-mediated tumor suppression. Taken together, our results showed that let-7a acted as a tumor suppressor in ES by targeting CDK6, and it may provide novel diagnostic and therapeutic options for human Ewing sarcoma clinical operation in future.
Alcohol dependence (AD) is a moderately heritable phenotype with a small number of known risk genes mapped via linkage or candidate gene studies. We considered 313 males from among 600 members of documented, extended pedigrees in which AD segregates collected in Northern Hunan Province, China. A joint analysis of both males and females could not be performed as the difference in alcohol consumption variance was too large. Genome-wide association analyses were performed for approximately 300,000 single nucleotide polymorphisms (SNPs). Significant associations found in the ALDH2 region for AD (minimum p = 4.73×10-8) and two AD-related phenotypes: flushing response (minimum p = 4.75×10-26) and maximum drinks in a 24-hour period (minimum p = 1.54×10-16). Association of previous candidate SNP, rs10774610 in CCDC63, was confirmed but resulted from linkage disequilibrium with ALDH2. ALDH2 is strongly associated with flushing response, AD, and maximum drinks in males, with nonsynonymous SNP rs671 explaining 29.2%, 7.9% and 22.9% of phenotypic variation, respectively, in this sample. When rs671 was considered as a candidate SNP in females, it explained 23.6% of the variation in flushing response, but alcohol consumption rates were too low among females – despite familial enrichment for AD – for an adequate test of association for either AD or maximum drinks. These results support a mediating effect of aldehyde dehydrogenase deficiency on alcohol consumption in males and a secondary, culturally-mediated limitation on alcohol consumption by females that should be appropriately modeled in future studies of alcohol consumption in populations where this may be a factor.
Alcohol Dependence; Maximum Drinks; Flushing Response; Genome-Wide Association; Aldehyde Dehydrogenase
In this study, a universal protein expression enhancement RNA tool, termed RNAe, was developed by modifying a recently discovered natural long non-coding RNA. At the moment, RNAe is the only technology for gene expression enhancement, as opposed to silencing, at the post-transcriptional level. With this technology, an expression enhancement of 50–1000% is achievable, with more than 200% enhancement achieved in most cases. This work identified the sufficient and necessary element for RNAe function, which was found to be merely 300 nucleotides long and was named minRNAe. It contains a 72-nt 5' pairing sequence which determines the specificity, a 167-nt short non-pairing interspersed nuclear element (SINE) B2 sequence which enhances ribosome recruitment to the target mRNA, and a poly(A) tail, provided together on a plasmid bearing the appropriate sequences. Cellular delivery of RNAe was achieved using routine transfection. The RNAe platform was validated in several widely-used mammalian cell lines. It was proven to be efficient and flexible in specifically enhancing the expression of various endogenous and exogenous proteins of diverse functions in a dose-dependent manner. Compared to the expression-inhibitory tool RNAi, the RNAe tool has a comparable effect size, with an enhancing as opposed to inhibitory effect. One may predict that this brand new technology for enhancing the production of proteins will find wide applications in both research and biopharmaceutical production.
The safety of herbal medicine products has been a widespread concern due to their complex chemical nature and lack of proper evaluation methods. We have adapted a sensitive and reproducible multiparametric cell-based high-content analysis assay to evaluate the hepatic-safety of four Chinese medicine injections and validated it with classical animal-based toxicity assays. Our results suggested that the reported hepatotoxicity by one of the drugs, Fufangkushen injection, could be attributed at least in part to the interference of mitochondrial function in human HepG2 cells by some of its constituents. This method should be useful for both preclinical screen in a drug discovery program and postclinical evaluation of herbal medicine preparations.
Tubulin cofactor A (TBCA), one of the members of tubulin cofactors, is of great importance in microtubule functions through participating in the folding of α/β-tubulin heterodimers in Saccharomyces cerevisiae. However, little is known about the roles of TBCA in filamentous fungi.
In this study, we characterized a TBCA orthologue FaTBCA in Fusarium asiaticum. The deletion of FaTBCA caused dramatically reduced mycelial growth and abnormal conidiation. The FaTBCA deletion mutant (ΔFaTBCA-3) showed increased sensitivity to low temperatures and even lost the ability of growth at 4°C. Microscopic observation found that hyphae of ΔFaTBCA-3 exhibited blebbing phenotypes after shifting from 25 to 4°C for 1- or 3-day incubation and approximately 72% enlarged nodes contained several nuclei after 3-day incubation at 4°C. However, hyphae of the wild type incubated at 4°C were phenotypically indistinguishable from those incubated at 25°C. These results indicate that FaTBCA is involved in cell division under cold stress (4°C) in F. asiaticum. Unexpectedly, ΔFaTBCA-3 did not exhibit increased sensitivity to the anti-microtubule drug carbendazim although quantitative real-time assays showed that the expression of FaTBCA was up-regulated after treatment with carbendazim. In addition, pathogenicity assays showed that ΔFaTBCA-3 exhibited decreased virulence on wheat head and on non-host tomato.
Taken together, results of this study indicate that FaTBCA plays crucial roles in vegetative growth, conidiation, temperature sensitivity and virulence in F. asiaticum.
Electronic supplementary material
The online version of this article (doi:10.1186/s12866-015-0374-z) contains supplementary material, which is available to authorized users.
TBCA; Conidiation; Cold sensitivity; F. asiaticum; Hyphal growth; Fusarium head blight