Pan-cancer transcriptomic data of ABI1 transcript variants and molecular constitutive elements identifies novel cancer metastatic and prognostic biomarkers.

BACKGROUND: Abelson interactor 1 (ABI1) is associated with the metastasis and prognosis of many malignancies. The association between ABI1 transcript spliced variants, their molecular constitutive exons and exon-exon junctions (EEJs) in 14 cancer types and clinical outcomes remains unsolved. OBJECTIVE: To identify novel cancer metastatic and prognostic biomarkers from ABI1 total mRNA, TSVs, and molecular constitutive elements. METHODS: Using data from TCGA and TSVdb database, the standard median of ABI1 total mRNA, TSV, exon, and EEJ expression was used as a cut-off value. Kaplan-Meier analysis, Chi-squared test (X2) and Kendall's tau statistic were used to identify novel metastatic and prognostic biomarkers, and Cox regression analysis was performed to screen and identify independent prognostic factors. RESULTS: A total of 35 ABI1-related factors were found to be closely related to the prognosis of eight candidate cancer types. A total of 14 ABI1 TSVs and molecular constitutive elements were identified as novel metastatic and prognostic biomarkers in four cancer types. A total of 13 ABI1 molecular constitutive elements were identified as independent prognostic biomarkers in six cancer types. CONCLUSIONS: In this study, we identified 14 ABI1-related novel metastatic and prognostic markers and 21 independent prognostic factors in total 8 candidate cancer types.

Identification of AP002498.1 and LINC01871 as prognostic biomarkers and therapeutic targets for distant metastasis of colorectal adenocarcinoma.

BACKGROUND: Increasing evidence suggests that lncRNA (Long non-coding RNA, lncRNA)-mediated ceRNA (competing endogenous RNA, ceRNA) networks are involved in the occurrence and progression of colorectal cancer (CRC). However, the roles of the lncRNA-miRNA-mRNA ceRNA network in distant metastasis of CRC are still unclear. METHODS: In this study, we constructed a specific ceRNA network to identify potential biomarkers and therapeutic targets for distant metastasis of CRC. Specifically, RNA-Seq data from The Cancer Genome Atlas (TCGA) were used to screen for differentially expressed lncRNAs (DElncRNAs) and mRNAs (DEmRNAs) related to metastasis. After validation and selection by qRT-PCR and univariate and multivariate analysis of the metastasis- and prognosis-related lncRNAs, the regulated microRNAs (miRNAs) and coexpressed mRNAs were used to construct a ceRNA network for distant metastasis of CRC. RESULTS: Two key distant metastasis-related DElncRNAs, AP002498.1 and LINC01871, were identified by univariate and multivariate analysis in combination with analyses of clinical data and expression levels. Furthermore, lncRNA-associated ceRNA subnetworks were constructed from the predicted miRNAs and 13 coexpressed DEmRNAs (SERPINA1, ITLN1, REG4, L1TD1, IGFALS, MUC5B, CIITA, CXCL9, CXCL10, GBP4, GNLY, IDO1, and NOS2). The AP002498.1- and LINC01871-associated ceRNA subnetworks regulated the expression of the target genes SERPINA1 and MUC5B and GNLY, respectively, through the associated miRNAs. CONCLUSION: The DElncRNA AP002498.1 and the LINC01871/miR-4644 and miR-185-5p/GNLY axes were identified as being closely associated with distant metastasis and could represent independent prognostic biomarkers or therapeutic targets in colorectal adenocarcinoma.

Histone H3 Acetylation Is Involved in Retinoid Acid-Induced Neural Differentiation through Increasing Mitochondrial Function.

Histone acetylation and mitochondrial function contribute importantly to neural differentiation, which is critically associated with neurodevelopmental disorders such as Down Syndrome (DS). However, whether and how histone acetylation regulates mitochondrial function and further affects neural differentiation has not been well described. In this study, when treated with retinoid acid (RA), the human neuroblastoma SH-SY5Y cell line was used as a neural differentiation model. We found that the acetylation of histone H3, especially H3 lysine 14 acetylation (H3K14ac), and mitochondrial function, including biogenesis and electron transport chain, were enhanced during neural differentiation. Specific inhibition of histone acetyltransferases (HATs) induced neural differentiation deficits, accompanied by downregulation of mitochondrial function. Furthermore, RA receptors (RARs) interacting with HATs were involved in the increased H3K14ac and the enhanced mitochondrial function during the neural differentiation process. Finally, receptor-interacting protein 140 (RIP140), a co-repressor of RARs, was also involved in regulating histone acetylation. RIP140 overexpression inhibited histone acetylation and mediated negative feedback on target genes which are involved in RA signaling. These findings evidenced that when interacting with RARs which had been negatively regulated by RIP140, RA promoted neural differentiation by promoting H3K14ac and enhanced mitochondrial function. This provides a molecular foundation for further investigations into abnormal neural development.

Phosphorylated Ser187-SNAP25-modulated hyperfunction of glutamatergic system in the vmPFC mediates depressive-like behaviors in male mice.

Dysfunctional glutamatergic neurotransmission contributes importantly to the pathophysiology of depression. However, the underlying neural mechanisms of glutamatergic dysfunction remain poorly understood. Here, we employed chronic unpredictable mild stress (CUMS) to induce depression-like behavior in male mice and to assess the alterations of glutamatergic system within the ventromedial prefrontal cortex (vmPFC). Male mice subjected to CUMS showed an increase in levels of glutamate content, synaptosomal GluN2B-NMDA receptors (GluN2B-NMDARs) and phosphorylated synaptosomal associated protein 25 KD of Ser187 (pSer187-SNAP25), which is involved in synaptic vesicular fusion processes in the vmPFC. Downregulation of pSer187-SNAP25 via the TAT-S187 fusion peptide efficiently alleviated CUMS-induced depressive-like behaviors in male mice by reversing the increase of glutamate content and synaptosomal GluN2B-NMDARs. These findings demonstrated a critical role for pSer187-SNAP25-mediated glutamatergic dysfunction in CUMS-induced depressive-like behaviors, suggesting the potential of pS187-SNAP25 inhibitors for further investigation on depression management.

Possible genetic cross-talk between Down syndrome and obstructive sleep apnea revealed by transcriptomic analysis.

PURPOSE: Down syndrome (DS) is linked to a higher prevalence of obstructive sleep apnea (OSA) than in the general population, which in turn contributes to worse cognitive impairment in DS. However, the shared pathogenic mechanisms for DS and OSA remain incompletely illustrated. This study was designed to decipher the genetic cross-talk between DS and OSA by bioinformatics approach. METHODS: Transcriptomic datasets of DS (GSE59630) and OSA (GSE135917) were accessed from the Gene Expression Omnibus (GEO) repository. After screening out the common differentially expressed genes (DEGs) for DS and OSA, gene ontology (GO) functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were carried out. A protein-protein interaction (PPI) network was then constructed to determine essential modules and hub genes. Finally, based on hub genes, transcriptional factor (TF)-gene interaction and TF-miRNA regulatory networks were constructed. RESULTS: DS and OSA showed 229 DEGs. Functional analyses revealed how oxidative stress and inflammatory response were critical in the progression of DS and OSA. Ten significant hub genes were identified, including TLR4, SOD1, IGF1, FGF2, NFE2L2, PECAM1, S100A8, S100A9, FCGR3A, and KCNA1, which were candidate targets for DS and OSA. CONCLUSIONS: We found that DS and OSA display similarities in their pathogenesis. Key genes and signaling pathways revealed to be in common between the two conditions could lead us to new therapeutic targets for DS and OSA.

Profiling Blautia at high taxonomic resolution reveals correlations with cognitive dysfunction in Chinese children with Down syndrome.

Introduction: Down syndrome (DS), the presence of a supernumerary chromosome 21, is associated with cognitive dysfunction caused by early neurodegenerative processes. Alterations in the gut microbiota were observed in Chinese children with DS, and the genus Blautia was associated with cognitive function in these children. Therefore, it is crucial to understand the detailed composition of this group at the species level and to explore the effect of specific species on cognitive function. Methods: In this study, Blautia-specific amplicon sequencing was conducted to identify the specific Blautia species in 15 children with DS and 15 matched healthy children. Results: The taxonomic analyses suggested that the Blautia taxa were clustered by disease status. The diversity of Blautia at the species level differed between DS patients and healthy controls, with the abundances of Blautia massiliensis and Blautia argi decreasing in DS children, while Blautia faecis was increased. Acetic acid, one of the metabolites of Blautia, was significantly reduced in the DS group. Of particular interest, Kyoto Encyclopaedia of Genes and Genomes analysis revealed decreased modules related to starch and sucrose metabolism and glycolysis. In addition, B. argi was positively related to DS cognitive scores, and B. faecis was negatively related to cognitive function, implying its role on the DS cognitive impairments. Discussion: Our study has important implications for understanding the important effects of specific species of Blautia on cognitive function and thus possibly provides a new strategy for future studies of cognitive improvement in individuals with DS.

Establishment of quantitative nested-PCR of Abelson interactor 1 transcript variant-11.

Abelson interactor 1 (ABI1), which presents 18 Transcript Variants (TSV), plays an important role in CRC metastasis. Different ABI1-TSVs play synergistic or antagonistic roles in the same pathophysiological events. ABI1 Transcript Variant-11 (ABI1-TSV-11) functionally promotes lymph node metastasis of left-sided colorectal cancer (LsCC) and is an independent molecular marker to evaluate the prognosis of patients with LsCC. However, there is still lack of a quick and accurate method to detect the expression of ABI-TSV-11, distinguishing ABI1-TSV-11 from other 17 TSVs. To establish a rapid method specific for ABI1-TSV-11detection, we developed a quantitative nested-PCR method composed of pre-amplification regular PCR using ABI1 universal primer pair and the followed Real Time (RT)-qPCR using ABI1-TSV-11 specific primer pair spanning exon-exon junction. ABI1-TSV-11-overexpressed SW480 and LoVo cell lines were used to verify the quantitative nested-PCR assay, and the sequencing data was used to evaluate the accuracy of ABI1-TSV-11 quantitative nested-PCR assay. The detection limit was 5.24×104 copies/ml. ABI1-TSV-11 quantitative nested-PCR provides a new technical means for the detection of ABI1-TSV-11.

Cell cycle arrest is an important mechanism of action of compound Kushen injection in the prevention of colorectal cancer.

Compound Kushen injection (CKI) is the most widely used traditional Chinese medicine preparation for the comprehensive treatment of colorectal cancer (CRC) in China, but its underlying molecular mechanisms of action are still unclear. The present study employed a network pharmacology approach, in which we constructed a "bioactive compound-target-pathway" network. Experimental RNA sequencing (RNA-Seq) analysis was performed to identify a key "bioactive compound-target-pathway" network for subsequent experimental validation. Cell cycle, proliferation, autophagy, and apoptosis assays and a model of azoxymethane/dextran sodium sulfate-induced colorectal carcinogenesis in mice were employed to detect the biological effect of CKI on CRC. Real-time reverse-transcription polymerase chain reaction, Western blot, and immunohistochemistry were performed to verify the selected targets and pathways. We constructed a predicted network that included 82 bioactive compounds, 34 targets, and 33 pathways and further screened an anti-CRC CKI "biological compound (hesperetin 7-O-rutinoside, genistein 7-O-rutinoside, and trifolirhizin)-target (p53 and checkpoint kinase 1 [CHEK1])" network that targeted the "cell cycle pathway". Validation experiments showed that CKI effectively induced the cell-cycle arrest of CRC cells in vitro and suppressed the development of CRC in vivo by downregulating the expression of p53 and CHEK1. Our findings confirmed that inducing cell-cycle arrest by CKI is an important mechanism of its anti-CRC action, which provides a direct and scientific experimental basis for the clinical application of CKI.

Altered gut microbiota correlates with cognitive impairment in Chinese children with Down's syndrome.

Down's syndrome (DS), a common chromosomal disease caused by chromosome 21 trisomy, is the main cause of cognitive impairment in children worldwide. Emerging evidence suggests that the microbiota-gut-brain axis plays a potential role in cognitive impairment. However, data regarding gut microbiota alterations in DS patients remain scarce, especially data from children with DS. This case-control study was conducted to explore the gut microbiota composition in Chinese DS children. Additionally, the potential association between gut microbiota and cognitive function in DS was evaluated. Microbiota communities in the feces of 15 DS subjects and 15 matched controls were investigated using high-throughput Illumina Miseq sequencing targeting the V3-V4 region of 16S rRNA gene. The relationships between gut microbiota composition and DS cognitive function scores were analyzed. The structure and richness of the gut microbiota differed between DS patients and healthy controls. The abundance of Acidaminococcaceae was decreased in DS patients. Moreover, the Kyoto Encyclopedia of Genes and Genomes analysis showed increased modules related to peptidases and pyrimidine metabolism. Overall, we confirmed that gut microbiota alterations occurred in Chinese patients with DS. Additionally, the fecal microbiota was closely related to DS cognitive impairment. Larger cohorts are needed to confirm these findings and to clarify the mechanisms involved. Elucidating these novel findings in the field of microbiota-gut-brain axis will provide a promising strategy for future studies of DS cognitive impairment.

Abelson interactor 1 splice isoform-L plays an anti-oncogenic role in colorectal carcinoma through interactions with WAVE2 and full-length Abelson interactor 1.

BACKGROUND: Expression of the full-length isoform of Abelson interactor 1 (ABI1), ABI1-p65, is increased in colorectal carcinoma (CRC) and is thought to be involved in one or more steps leading to tumor progression or metastasis. The ABI1 splice isoform-L (ABI1-SiL) has conserved WAVE2-binding and SH3 domains, lacks the homeo-domain homologous region, and is missing the majority of PxxP- and Pro-rich domains found in full-length ABI1-p65. Thus, ABI1-SiL domain structure suggests that the protein may regulate CRC cell morphology, adhesion, migration, and metastasis via interactions with the WAVE2 complex pathway. AIM: To investigate the potential role and underlying mechanisms associated with ABI1-SiL-mediated regulation of CRC. METHODS: ABI1-SiL mRNA expression in CC tissue and cell lines was measured using both qualitative reverse transcriptase-polymerase chain reaction (RT-PCR) and real-time quantitative RT-PCR. A stably ABI1-SiL overexpressing SW480 cell model was constructed using Lipofectamine 2000, and cells selected with G418. Image J software, CCK8, and transwell assays were used to investigate SW480 cell surface area, proliferation, migration, and invasion. Immunoprecipitation, Western blot, and co-localization assays were performed to explore intermolecular interactions between ABI1-SiL, WAVE2, and ABI1-p65 proteins. RESULTS: ABI1-SiL was expressed in normal colon tissue and was significantly decreased in CRC cell lines and tissues. Overexpression of ABI1-SiL in SW480 cells significantly increased the cell surface area and inhibited the adhesive and migration properties of the cells, but did not alter their invasive capacity. Similar to ABI1-p65, ABI1-SiL still binds WAVE2, and the ABI1-p65 isoform in SW480 cells. Furthermore, co-localization assays confirmed these intermolecular interactions. CONCLUSION: These results support a model in which ABI1-SiL plays an anti-oncogenic role by competitively binding to WAVE2 and directly interacting with phosphorylated and non-phosphorylated ABI1-p65, functioning as a dominant-negative form of ABI1-p65.

The roles and prognostic significance of ABI1-TSV-11 expression in patients with left-sided colorectal cancer.

Abnormally expressed and/or phosphorylated Abelson interactor 1 (ABI1) participates in the metastasis and progression of colorectal cancer (CRC). ABI1 presents as at least 12 transcript variants (TSVs) by mRNA alternative splicing, but it is unknown which of them is involved in CRC metastasis and prognosis. Here, we firstly identified ABI1-TSV-11 as a key TSV affecting the metastasis and prognosis of left-sided colorectal cancer (LsCC) and its elevated expression is related to lymph node metastasis and shorter overall survival (OS) in LsCC by analyzing data from The Cancer Genome Atlas and TSVdb. Secondly, ABI1-TSV-11 overexpression promoted LoVo and SW480 cells adhesion and migration in vitro, and accelerated LoVo and SW480 cells lung metastasis in vivo. Finally, mechanism investigations revealed that ABI1-isoform-11 interacted with epidermal growth factor receptor pathway substrate 8 (ESP8) and regulated actin dynamics to affect LoVo and SW480 cells biological behaviors. Taken together, our data demonstrated that ABI1-TSV-11 plays an oncogenic role in LsCC, it is an independent risk factor of prognosis and may be a potential molecular marker and therapeutic target in LsCC.

Comprehensive analysis of ceRNA networks reveals prognostic lncRNAs related to immune infiltration in colorectal cancer.

BACKGROUND: Competing endogenous RNA (ceRNA) represents a class of RNAs (e.g., long noncoding RNAs [lncRNAs]) with microRNA (miRNA) binding sites, which can competitively bind miRNA and inhibit its regulation of target genes. Increasing evidence has underscored the involvement of dysregulated ceRNA networks in the occurrence and progression of colorectal cancer (CRC). The purpose of this study was to construct a ceRNA network related to the prognosis of CRC and further explore the potential mechanisms that affect this prognosis. METHODS: RNA-Seq and miRNA-Seq data from The Cancer Genome Atlas (TCGA) were used to identify differentially expressed lncRNAs (DElncRNAs), microRNAs (DEmiRNAs), and mRNAs (DEmRNAs), and a prognosis-related ceRNA network was constructed based on DElncRNA survival analysis. Subsequently, pathway enrichment, Pearson correlation, and Gene Set Enrichment Analysis (GSEA) were performed to determine the function of the genes in the ceRNA network. Gene Expression Profiling Interactive Analysis (GEPIA) and immunohistochemistry (IHC) were also used to validate differential gene expression. Finally, the correlation between lncRNA and immune cell infiltration in the tumor microenvironment was evaluated based on the CIBERSORT algorithm. RESULTS: A prognostic ceRNA network was constructed with eleven key survival-related DElncRNAs (MIR4435-2HG, NKILA, AFAP1-AS1, ELFN1-AS1, AC005520.2, AC245884.8, AL354836.1, AL355987.4, AL591845.1, LINC02038, and AC104823.1), 54 DEmiRNAs, and 308 DEmRNAs. The MIR4435-2HG- and ELFN1-AS1-associated ceRNA subnetworks affected and regulated the expression of the COL5A2, LOX, OSBPL3, PLAU, VCAN, SRM, and E2F1 target genes and were found to be related to prognosis and tumor-infiltrating immune cell types. CONCLUSIONS: MIR4435-2HG and ELFN1-AS1 are associated with prognosis and tumor-infiltrating immune cell types and could represent potential prognostic biomarkers or therapeutic targets in colorectal carcinoma.

Cognitive impairments in adult mice with RIP140 overexpression in neural stem cells.

Receptor-interacting protein 140 (RIP140) is a transcription co-regulator of several transcription factors and a signal transduction regulator. RIP140 was recently implicated in the regulation of cognitive functions. The gene that encodes RIP140 is located on chromosome 21. An increase in RIP140 expression was observed in the fetal cerebral cortex and hippocampus in Down syndrome patients who exhibited strong cognitive disabilities. We hypothesized that RIP140 overexpression affects cognitive function in adult neural development. The present study used a Cre-dependent adeno-associated virus to selectively overexpress RIP140 in neural stem cells using nestin-Cre mice. RIP140 overexpression efficiency was evaluated at the subgranular zone (SGZ) of the dorsal dentate gyrus (dDG) and the subventricular zone (SVZ) of the lateral ventricles (LVs). Mice with RIP140 overexpression in the SGZ exhibited deficits in cognitive function and spatial learning and memory, measured in the Morris water maze, object-place recognition test, and novel object recognition test. However, overexpression of RIP140 in SVZ only impaired performance in the Morris water maze and novel object recognition test but not in the object-place recognition test. Altogether, these results indicated defects in cognitive functions that were associated with RIP140 overexpression in neural stem cells and revealed a behavioral phenotype that may be used as a framework for further investigating the neuropathogenesis of Down syndrome.

RIP140/PGC-1α axis involved in vitamin A-induced neural differentiation by increasing mitochondrial function.

Vitamin A deficiency and mitochondrial dysfunction are both associated with neural differentiation-related disorders, such as Alzheimer's disease (AD) and Down syndrome (DS). The mechanism of vitamin A-induced neural differentiation and the notion that vitamin A can regulate the morphology and function of mitochondria in its induction of neural differentiation through the RIP140/PGC-1α axis are unclear. The aim of this study was to investigate the roles and underlying mechanisms of RIP140/PGC-1α axis in vitamin A-induced neural differentiation. Human neuroblastoma cells (SH-SY5Y) were used as a model of neural stem cells, which were incubated with DMSO, 9-cis-retinoic acid (9-cis-RA), 13-cis-retinoic acid (13-cis-RA) and all-trans-retinoic acid (at-RA). Neural differentiation of SH-SY5Y was evaluated by Sandquist calculation, combined with immunofluorescence and real-time polymerase chain reaction (PCR) of neural markers. Mitochondrial function was estimated by ultrastructure assay using transmission electron microscopy (TEM) combined with the expression of PGC-1α and NEMGs using real-time PCR. The participation of the RA signaling pathway was demonstrated by adding RA receptor antagonists. Vitamin A derivatives are able to regulate mitochondrial morphology and function, and furthermore to induce neural differentiation through the RA signaling pathway. The RIP140/PGC-1α axis is involved in the regulation of mitochondrial function in vitamin A derivative-induced neural differentiation.

Correlation between receptor-interacting protein 140 expression and directed differentiation of human embryonic stem cells into neural stem cells.

Overexpression of receptor-interacting protein 140 (RIP140) promotes neuronal differentiation of N2a cells via extracellular regulated kinase 1/2 (ERK1/2) signaling. However, involvement of RIP140 in human neural differentiation remains unclear. We found both RIP140 and ERK1/2 expression increased during neural differentiation of H1 human embryonic stem cells. Moreover, RIP140 negatively correlated with stem cell markers Oct4 and Sox2 during early stages of neural differentiation, and positively correlated with the neural stem cell marker Nestin during later stages. Thus, ERK1/2 signaling may provide the molecular mechanism by which RIP140 takes part in neural differentiation to eventually affect the number of neurons produced.

Receptor-interacting protein 140 overexpression promotes neuro-2a neuronal differentiation by ERK1/2 signaling.

BACKGROUND: Abnormal neuronal differentiation plays an important role in central nervous system (CNS) development abnormalities such as Down syndrome (DS), a disorder that results directly from overexpression of genes in trisomic cells. Receptor-interacting protein 140 (RIP140) is significantly upregulated in DS brains, suggesting its involvement in DS CNS development abnormalities. However, the role of RIP140 in neuronal differentiation is still not clear. The current study aimed to investigate the effect of RIP140 overexpression on the differentiation of neuro-2a (N2a) neuroblastoma cells, in vitro. METHODS: Stably RIP140-overexpressing N2a (N2a-RIP140) cells were used as a neurodevelopmental model, and were constructed by lipofection and overexpression validated by real-time polymerase chain reaction and Western blot. Retinoic acid (RA) was used to stimulate N2a differentiation. Combining the expression of Tuj1 at the mRNA and protein levels, the percentage of cells baring neurites, and the number of neurites per cell body was semi-quantified to determine the effect of RIP140 on differentiation of N2a cells. Furthermore, western blot and the ERK1/2 inhibitor U0126 were used to identify the specific signaling pathway by which RIP140 induces differentiation of N2a cells. Statistical significance of the differences between groups was determined by one-way analysis of variance followed by the Dunnett test. RESULTS: Compared to untransfected N2a cells RIPl40 expression in N2a-RIP140 cells was remarkably upregulated at both the mRNA and protein levels. N2a-RIP140 cells had a significantly increased percentage of cells baring neurites, and numbers of neurites per cell, as compared to N2a cells, in the absence and presence of RA (P < 0.05). In addition, Tuj1, a neuronal biomarker, was strongly upregulated in N2a-RIP140 cells (P < 0.05) and phosphorylated ERK1/2 (p-ERK1/2) levels in N2a-RIP140 cells were dramatically increased, while differentiation was inhibited by the ERK1/2-specific inhibitor U0126. CONCLUSIONS: RIP140 overexpression promotes N2a cell neuronal differentiation by activating the ERK1/2 pathway.

G-protein coupled receptor 34 activates Erk and phosphatidylinositol 3-kinase/Akt pathways and functions as alternative pathway to mediate p185Bcr-Abl-induced transformation and leukemogenesis.

Tyrosine 177 and the Src homology 2 (SH2) domain play important roles in linking p185Bcr-Abl to downstream pathways critical for cell growth and survival. However, a mutant p185(Y177FR552L) (p185(YR)), in which tyrosine 177 and arginine 552 in the SH2 domain are mutated, is still capable of transforming hematopoietic cells in vitro. Transplant of these cells into syngeneic mice also leads to leukemogenesis, albeit with a phenotype distinct from that produced by wild-type p185Bcr-Abl (p185(wt))-transformed cells. Here we show that G-protein coupled receptor 34 (Gpr34) expression is markedly up-regulated in p185(YR)-transformed cells compared to those transformed by p185(wt). Knockdown of Gpr34 in p185(YR) cells is sufficient to suppress growth factor-independent proliferation and survival in vitro and attenuate leukemogenesis in vivo. The Erk and phosphatidylinositol 3-kinase/Akt pathways are activated in p185(YR) cells and the activation is dependent on Gpr34 expression. These studies identify Gpr34 as an alternative pathway that may mediate p185Bcr-Abl-induced transformation and leukemogenesis.

[Effects of receptor-interacting protein 140 gene over-expression on the migration and proliferation of neuroblastoma cells].

OBJECTIVE: To explore the effects of receptor interacting protein (RIP)140 gene over-expression upon the migration and proliferation of neuroblastoma cells in vitro. METHODS: The N2a RIP140 over-expression model (N2a-rip140) was constructed by lipofection and validated by real-time polymerase chain reaction (PCR) and Western blot. The migration and proliferation capabilities were compared between N2a-rip140 and its parents by Transwell chamber and CCK-8. RESULTS: The N2a RIP140 over-expression model (N2a-rip140) was successfully constructed. Compared to N2a group, the RIP140 mRNA and protein expression levels of N2a-rip140 were remarkably up-regulated. Transwell assay showed that the over-expression of RIP140 inhibited N2a cell migration ((70 ± 20) vs (5 ± 4) cells, P < 0.05). CCK-8 assay showed that the proliferation rates of N2a group at 24, 48, 72 h were 1.567 ± 0.107, 3.018 ± 0.212 and 4.112 ± 0.221 respectively. And those of N2a-rip140 group were 1.561 ± 0.281, 2.232 ± 0.235 and 4.025 ± 0.217 respectively. No significant difference existed in proliferation rates at different timepoints among N2a, N2a-M and N2a-rip140 groups (all P > 0.05). CONCLUSION: RIP140 over-expression effectively inhibits N2a cell migration. However it has no significant effect on the proliferation of N2a cells.

Altered methylation of IGF2 DMR0 is associated with neural tube defects.

Neural tube defects (NTDs) are serious congenital malformation of fusion failure of the neural tube during early embryogenesis. DNA methylation disorders have been found in NTD-affected fetuses, and are correlated to the risk of NTDs. The insulin-like growth factor 2 (IGF2) gene, maternally imprinted, has a key role in fetal development. IGF2 transcription is partly controlled by differentially methylated regions (DMRs) 0 and 2. To assess whether disturbed methylation pattern increases the incidence of NTDs, we employed matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) to quantify CpG methylation levels of DMR2 and 0 in fetuses with or without NTDs. We found that the methylation level of IGF2 DMR0 increased significantly in the brain tissues of NTD-affected fetuses. And hypermethylation of DMR0 was associated with an increased risk of NTDs, with an odds ratio of 5.375 (95 % CI: 1.447-19.965; p = 0.007). IGF2 mRNA expression was negatively correlated with the methylation level of DMR0 (R (2) = 0.893; p = 0.000) in HCT15 cells. These results highlights that IGF2 DMR0 hypermethylation is a potential risk factor of NTD, and IGF2 gene is a promising candidate gene to study for a greater understanding of the cause of NTDs.

[Effects of receptor interacting protein 140 on the biological activity of glia cells].

OBJECTIVE: To explore the effects of receptor interacting protein (RIP) 140 gene overexpression upon the in vitro proliferation, apoptosis, invasion and migration of microglioma cells. METHODS: The BV-2 RIP140 overexpression model (BV-2-1) was constructed by Lipofection and G418 selection, then validated by real-time PCR and Western blotting. The proliferation, apoptosis, invasion and migration potencies were compared between BV-2-1 and its parents by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium (MTT) assay, flow cytometry and Transwell chamber. RESULTS: The BV-2-1 model was successfully constructed. Compared to those of the BV-2 group, the RIP140 mRNA and protein expression levels of BV-2-1 were markedly higher than those of the BV-2 group (t = 49.794, P < 0.01). MTT assay showed that the absorbance values in the BV-2 group were 1.157 ± 0.013, 1.679 ± 0.005 and 2.609 ± 0.008 at 24, 48, and 72 hours respectively. And those were 0.929 ± 0.013, 1.188 ± 0.008 and 1.528 ± 0.012 in the BV-2-1 group respectively. The proliferation at the time points of 48 and 72 hours of the BV-2-1 group were significantly lower than that of the BV-2 group (t = 6.058 and 9.245, both P < 0.01). Annexin-V staining showed that there were significant differences in the apoptosis rates between the BV-2 and BV-2-1 cells [(5.35 ± 0.23)% vs (3.46 ± 0.45)%, t = 6.619, P = 0.003)]. Transwell assay showed that the invaded cell number of the BV-2-1 group was 166 ± 43. And it was obviously higher than that of the BV-2 group (93 ± 32, t = 3.403, P = 0.007). Transwell assay also showed that the migrated cell number of BV-2 cells was 101 ± 25. And the migration potency of the BV-2-1 group (202 ± 50) was significantly stronger than that of the BV-2 group (t = 4.104, P = 0.002). CONCLUSION: RIP140 effectively inhibits the proliferation and facilitates the apoptosis of microglioma cells. And it may effectively facilitate the in vitro invasion and migration of microglioma cells.