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To study the clinical value of HEIH hyperexpression in gastric cancer and the molecular mechanism of promoting malignant proliferation of gastric cancer cells, qRT-PCR was used to detect the expression of HEIH in gastric cancer and nontumor gastric tissues. HEIH interference sequence was constructed to downregulate HEIH expression in MGC-803 and BGC-823 cell lines. CCK8, clonogenesis, and Transwell assay were used to detect the effects of HEIH on proliferation and invasion of tumor cells. The protein levels of STAT3, p-STAT3, P62, and LC3 were detected by Western blotting. The results showed that HEIH was highly expressed in gastric cancer ( P < 0.01 ). Interference of HEIH expression in MGC-803 and BGC-823 cells reduced the proliferation and invasion of gastric cancer cells, and the results were statistically significant ( P < 0.05 ). HEIH acts as a miRNA sponge for miR-4500. HEIH promotes gastric cancer development by inhibiting miR-4500. STAT3 is a downstream target of miR-4500. HEIH inhibits autophagy and promotes glycolysis. In conclusion, HEIH is highly expressed in gastric cancers. HEIH promotes malignant proliferation and development of gastric cancer cells. HEIH may be a new candidate site for pathological diagnosis and molecular drug therapy for future clinical treatment of gastric cancer.

The human transcriptome contains many types of noncoding RNAs, which rival the number of protein-coding species. From long noncoding RNAs (lncRNAs) that are over 200 nucleotides long to piwi-interacting RNAs (piRNAs) of only 20 nucleotides, noncoding RNAs play important roles in regulating transcription, epigenetic modifications, translation, and cell signaling. Roles for noncoding RNAs in disease mechanisms are also being uncovered, and several species have been identified as potential drug targets. On May 11-14, 2021, the Keystone eSymposium "Noncoding RNAs: Biology and Applications" brought together researchers working in RNA biology, structure, and technologies to accelerate both the understanding of RNA basic biology and the translation of those findings into clinical applications.

Congestive heart failure (HF) is a known risk factor of contrast-induced acute kidney injury (CI-AKI). However, the relationship of the classification and severity of HF with CI-AKI remains under-explored. From January 2009 to April 2019, we recruited patients undergoing elective PCI who had complete pre- and post-operative creatinine data. According to the levels of ejection fraction (EF), HF was classified as HF with reduced EF (HFrEF) [EF p value p value p value p value = 0.031). Additionally, lower levels of EF were risk factors for CI-AKI in the HFrEF and HFmrEF groups, but not in the HFpEF group. NT-proBNP was an independent risk factor for CI-AKI in the HFrEF, HFmrEF and HFpEF groups. Elevated levels of NT-proBNP are independent risk factors for CI-AKI irrespective of the classification of HF. Lower levels of EF were risk factors for CI-AKI in the HFrEF and HFmrEF groups, but not in the HFpEF group.

Stable propagation of epigenetic information is important for maintaining cell identity in multicellular organisms. However, it remains largely unknown how mono-ubiquitinated histone H2A on lysine 119 (H2AK119ub1) is established and stably propagated during cell division. In this study, we found that the proteins RYBP and YAF2 each specifically bind H2AK119ub1 to recruit the RYBP-PRC1 or YAF2-PRC1 complex to catalyse the ubiquitination of H2A on neighbouring nucleosomes through a positive-feedback model. Additionally, we demonstrated that histone H1-compacted chromatin enhances the distal propagation of H2AK119ub1, thereby reinforcing the inheritance of H2AK119ub1 during cell division. Moreover, we showed that either disruption of RYBP/YAF2-PRC1 activity or impairment of histone H1-dependent chromatin compaction resulted in a significant defect of the maintenance of H2AK119ub1. Therefore, our results suggest that histone H1-dependent chromatin compaction plays a critical role in the stable propagation of H2AK119ub1 by RYBP/YAF2-PRC1 during cell division.

Long noncoding RNAs (lncRNAs) and promoter- or enhancer-associated unstable transcripts locate preferentially to chromatin, where some regulate chromatin structure, transcription and RNA processing1-13. Although several RNA sequences responsible for nuclear localization have been identified-such as repeats in the lncRNA Xist and Alu-like elements in long RNAs14-16-how lncRNAs as a class are enriched at chromatin remains unknown. Here we describe a random, mutagenesis-coupled, high-throughput method that we name 'RNA elements for subcellular localization by sequencing' (mutREL-seq). Using this method, we discovered an RNA motif that recognizes the U1 small nuclear ribonucleoprotein (snRNP) and is essential for the localization of reporter RNAs to chromatin. Across the genome, chromatin-bound lncRNAs are enriched with 5' splice sites and depleted of 3' splice sites, and exhibit high levels of U1 snRNA binding compared with cytoplasm-localized messenger RNAs. Acute depletion of U1 snRNA or of the U1 snRNP protein component SNRNP70 markedly reduces the chromatin association of hundreds of lncRNAs and unstable transcripts, without altering the overall transcription rate in cells. In addition, rapid degradation of SNRNP70 reduces the localization of both nascent and polyadenylated lncRNA transcripts to chromatin, and disrupts the nuclear and genome-wide localization of the lncRNA Malat1. Moreover, U1 snRNP interacts with transcriptionally engaged RNA polymerase II. These results show that U1 snRNP acts widely to tether and mobilize lncRNAs to chromatin in a transcription-dependent manner. Our findings have uncovered a previously unknown role of U1 snRNP beyond the processing of precursor mRNA, and provide molecular insight into how lncRNAs are recruited to regulatory sites to carry out chromatin-associated functions.

Objectives Literature on dermatology outpatient demographic and clinical data is limited, and the few studies on this topic are mainly conducted overseas, with medical systems and case mix different to Australia. This study presents demographic data relating to dermatology public outpatient referrals to a tertiary hospital in Brisbane, Australia, and determines what additional structured data should be collected to formulate and evaluate initiatives to address service issues such as referral quality, triage process and wait times. Methods A four-year retrospective audit was undertaken, summarising all referrals (n = 7140) and clinical dermatology encounters (n = 53 844) between January 2016 and December 2019 at Princess Alexandra Hospital (PAH), the largest hospital in Metro South Health (MSH), serving a population of one million. PAH has one of the two largest public dermatology clinics in Queensland and is the only dermatology service within MSH. Results Patient demographic data, wait time by triage category, referral rates over time and encounter durations were collected. Structured diagnostic data (e.g. ICD-10 coding) of the provisional diagnosis, comorbidities, medications and the final diagnosis are not collected in a structured format and would be a valuable addition. Conclusions The clinical burden of public dermatology is increasing. Both collection and analysis of structured data pertaining to the referrals and encounters are important to help formulate, implement and evaluate initiatives that aim to improve health service provision in this area.

Background Renal ischemia/reperfusion (I/R) injury (RI/RI) is a common complication of diabetes mellitus (DM) in surgical practice. Oxidative stress plays a crucial role in this process. Recombinant human erythropoietin (rhEPO) is usually used to treat anemia resulting from several diseases. However, the functional involvement of rhEPO in diabetic RI/RI remains unclear. The present study was intended to investigate the antioxidant role of rhEPO on RI/RI in DM rats. Methods The bilateral renal arteries and veins of streptozotocin-induced diabetic rats were subjected to 45 minutes of ischemia followed by 1, 6, and 24 hours of reperfusion with or without rhEPO pretreatment at the beginning of an I/R procedure. The renal tissue pathomorphology, renal function, oxidative stress, and inflammatory response were evaluated by detection of a series of indices by hematoxylin-eosin staining, commercial kits, enzyme-linked immunosorbent assay, and spectrophotofluorometry, respectively. Results Compared to the I/R group, renal function was significantly advanced in the erythropoietin group, whose subjects were also subjected to renal tissue injury, oxidative stress, and inflammation. Conclusion These results suggest that rhEPO preconditioning can attenuate diabetic RI/RI through regulating endogenous antioxidant activity.

The aim of this study is to assess the quality and reliability of online videos on ankylosing spondylitis (AS) exercises from the five most popular video websites in China.Cross-sectional and descriptive study.We searched the video websites of Youku, Tencent, Tudou, IQiYi, and bilibili on February 15, 2020, using the keywords "Ankylosing spondylitis exercise" "Ankylosing spondylitis rehabilitation" and "Ankylosing spondylitis therapy" A total of 114 videos were included in the study and evaluated according to the Global Quality Scale (GQS) and modified DISCERN tool.According to the GQS, the videos were classified as high quality (12.3%, n = 14), intermediate quality (63.2%, n = 72), and low quality (24.6%, n = 28). Using the modified DISCERN tool, the videos were divided into useless (53.5%, n = 61), useful (35.1%, n = 40), and misleading (11.4%, n = 13).The analysis shows that the quality and reliability of online videos related to exercise therapy for Ankylosing spondylitis (AS) should be improved and supervised in China. Hospitals, universities, and medical doctors should make more useful and high-quality videos to provide effective exercise guidance for AS patients.ZIEL: Ziel der hier vorliegenden Studie war es, die Qualität und Reliabilität von Online-Videos zu Bewegungsübungen bei ankylosierender Spondylitis (AS) von den 5 in China am meisten verbreiteten Video-Websites zu beurteilen.Es handelt sich um eine deskriptive Studie im Querschnittdesign.Die Autoren durchsuchten die Video-Websites von Youku, Tencent, Tudou und IQiYi sowie Bilibili am 15. Februar 2020 unter Verwendung der Schlüsselwörter „ankylosing spondylitis exercise“, „ankylosing spondylitis rehabilitation“ und „ankylosing spondylitis therapy“. Es wurden 114 Videos in die Studie einbezogen und anhand der Global Quality Scale (GQS) sowie mit dem modifizierten DISCERN-Fragebogen beurteilt.Gemäß GQS wurden die Videos als von hoher Qualität (12,3%; n = 14), mittlerer Qualität (63,2%; n = 72) und geringer Qualität (24,6%; n = 28) eingestuft. Anhand des DISCERN-Fragebogens wurden die Videos in nutzlos (53,5%; n = 61), nützlich (35,1%; n = 40) und irreführend (11,4%; n = 13) eingeteilt.Die Auswertung zeigt, dass in China die Qualität und Reliabilität von Online-Videos zur Bewegungstherapie bei AS verbessert und überprüft werden sollten. Krankenhäuser, Universitäten und Ärzte sollten Videos mit größerem Nutzen und von hoher Qualität entwickeln, um so eine effektive Übungsanleitung für AS-Patienten zur Verfügung zu stellen.

RNA and RNA-binding proteins (RBPs) control multiple biological processes. The spatial and temporal arrangement of RNAs and RBPs underlies the delicate regulation of these processes. The strategy called CLIP-seq (cross-linking and immunoprecipitation) has been developed to capture endogenous protein-RNA interactions with UV cross-linking followed by immunoprecipitation. Despite the wide use of conventional CLIP-seq method in RBP study, the CLIP experiment is limited by the availability of the high-quality antibodies, potential contaminants from the co-purified RBPs, requirement of isotope manipulation, and potential loss of information during tedious experimental procedure. Here we described a modified CLIP-seq method called

The subcellular localization of RNAs is regulated by cis-regulatory elements together with interacting trans factors. Here we describe a high-throughput sequencing-based method named REL-seq (RNA elements for subcellular localization by sequencing) to identify the cis-elements that contribute to RNA subcellular localization. By coupling REL-seq with random mutagenesis (mutREL-seq), we can further narrow down the cis-elements to key motifs at single-nucleotide resolution.

RNA and RNA-binding proteins (RBPs) control multiple biological processes. The spatial and temporal arrangement of RNAs and RBPs underlies the delicate regulation of these processes. A strategy called CLIP-seq (cross-linking and immunoprecipitation) has been developed to capture endogenous protein-RNA interactions with UV cross-linking followed by immunoprecipitation. Despite the wide use of conventional CLIP-seq method in RBP study, the CLIP method is limited by the availability of high-quality antibodies, potential contaminants from the copurified RBPs, requirement of isotope manipulation, and potential loss of information during a tedious experimental procedure. Here we describe a modified CLIP-seq method called FbioCLIP-seq using the FLAG-biotin tag tandem purification. Through tandem purification and stringent wash conditions, almost all the interacting RNA-binding proteins are removed. Thus, the RNAs interacting indirectly mediated by these copurified RBPs are also reduced. Our FbioCLIP-seq method allows efficient detection of direct protein-bound RNAs without SDS-PAGE and membrane transfer procedures in an isotope-free and protein-specific antibody-free manner.

There is an increasing interest in understanding how three-dimensional organization of the genome is regulated. Different strategies have been used to identify genome-wide chromatin interactions. However, owing to current limitations in resolving genomic contacts, visualization and validation of these genomic loci at subkilobase resolution remain unsolved to date. Here, we describe Tn5 transposase-based fluorescence in situ hybridization (Tn5-FISH), a polymerase chain reaction-based, cost-effective imaging method, which can colocalize the genomic loci at subkilobase resolution, dissect genome architecture, and verify chromatin interactions detected by chromatin configuration capture-derived methods. To validate this method, short-range interactions in the keratin-encoding gene (KRT) locus in the topologically associated domain were imaged by triple-color Tn5-FISH, indicating that Tn5-FISH is very useful to verify short-range chromatin interactions inside the contact domain and TAD. Therefore, Tn5-FISH can be a powerful molecular tool for clinical detection of cytogenetic changes in numerous genetic diseases such as cancers.

Summary: Repetitive elements are abundantly distributed in mammalian genomes. Here, we reveal a striking association between repeat subtypes and gene function. SINE, L1, and low-complexity repeats demarcate distinct functional categories of genes and may dictate the time and level of gene expression by providing binding sites for different regulatory proteins. Importantly, imaging and sequencing analysis show that L1 repeats sequester a large set of genes with specialized functions in nucleolus- and lamina-associated inactive domains that are depleted of SINE repeats. In addition, L1 transcripts bind extensively to its DNA in embryonic stem cells (ESCs). Depletion of L1 RNA in ESCs leads to relocation of L1-enriched chromosomal segments from inactive domains to the nuclear interior and de-repression of L1-associated genes. These results demonstrate a role of L1 DNA and RNA in gene silencing and suggest a general theme of genomic repeats in orchestrating the function, regulation, and expression of their host genes. : Lu et al. report a striking association between genomic repeats and gene regulation and demonstrate a key role of L1 repeat RNA in sequestering L1-rich sequences and associated genes in inactive domains for silencing, revealing a general theme of repeat sequences in shaping gene regulatory networks within their host genome. Keywords: repetitive elements, L1, B1 or Alu, SINE, low-complexity repeats, gene function, regulation and expression, nucleolus and lamina-associated domains, L1 RNA, gene silencing

Exploration and dissection of potential actions and effects of long noncoding RNA (lncRNA) in animals remain challenging. Here, using multiple knockout mouse models and single cell RNA sequencing, we demonstrate that the divergent lncRNA

RNA-binding proteins (RBPs) play pivotal roles in directing RNA fate and function. Yet the current annotation of RBPs is largely limited to proteins carrying known RNA-binding domains. To systematically reveal dynamic RNA–protein interactions, we surveyed the human proteome by a protein array-based approach and identified 671 proteins with RNA-binding activity. Among these proteins, 525 lack annotated RNA-binding domains and are enriched in transcriptional and epigenetic regulators, metabolic enzymes, and small GTPases. Using an improved CLIP (crosslinking and immunoprecipitation) method, we performed genome-wide target profiling of isocitrate dehydrogenase 1 (IDH1), a novel RBP. IDH1 binds to thousands of RNA transcripts with enriched functions in transcription and chromatin regulation, cell cycle and RNA processing. Purified IDH1, but not an oncogenic mutant, binds directly to GA- or AU-rich RNA that are also enriched in IDH1 CLIP targets. Our study provides useful resources of unconventional RBPs and IDH1-bound transcriptome, and convincingly illustrates, for the first time, the in vivo and in vitro RNA targets and binding preferences of IDH1, revealing an unanticipated complexity of RNA regulation in diverse cellular processes.

Chronic intestinal inflammation is closely associated with colon cancer development and STAT3 seems to take center stage in bridging chronic inflammation to colon cancer progress. Here, we discovered that DICER1 was significantly downregulated in response to IL-6 or LPS stimulation and identified a novel mechanism for DICER1 downregulation via proteasomal degradation by ubiquitin ligase complex of CUL4A(DCAF1) in colon cancer cells. Meanwhile, PI3K-AKT signaling pathway phosphorylated DICER1 and contributed to its proteasomal degradation. The regulation of DICER1 by CUL4A(DCAF1) affected cell growth and apoptosis which is controlled by IL-6 activated Jak-STAT3 pathway. Intervention of CUL4A(DCAF1) ubiquitin ligase complex led to fluctuation in expression levels of DICER1 and microRNAs, and thus affected tumor growth in a mouse xenograft model. A panel of microRNAs that were downregulated by IL-6 stimulation was rescued by siRNA-CUL4A, and their predicated functions are involved in regulation of cell proliferation, apoptosis and motility. Furthermore, clinical specimen analysis revealed that decreased DICER1 expression was negatively correlated with STAT3 activation and cancer progression in human colon cancers. DICER1 and p-STAT3 expression levels correlated with 5-year overall survival of colon cancer patients. Consequently, this study proposes that inflammation-induced Jak-STAT3 signaling leads to colon cancer development through proteasomal degradation of DICER1 by ubiquitin ligase complex of CUL4A(DCAF1), which suggests a novel therapeutic opportunity for colon cancer.

Summary: Embryonic stem cells (ESCs) exhibit high levels of ribosomal RNA (rRNA) transcription and ribosome biogenesis. Here, we reveal an unexpected role for an essential DEAD-box helicase, DDX18, in antagonizing the polycomb repressive complex 2 (PRC2) to prevent deposition of the repressive H3K27me3 mark onto rDNA in pluripotent cells. DDX18 binds and sequesters PRC2 in the outer layer of the nucleolus and counteracts PRC2 complex formation in vivo and in vitro. DDX18 knockdown leads to increased occupancy of PRC2 and H3K27me3 at rDNA loci, accompanied by drastically decreased rRNA transcription and reduced ribosomal protein expression and translation. Auxin-induced rapid degradation of DDX18 enhances PRC2 binding at rDNA. The inhibition of PRC2 partially rescues the effects of DDX18 depletion on rRNA transcription and ESC self-renewal. These results demonstrate a critical role for DDX18 in safeguarding the chromatin and transcriptional integrity of rDNA by counteracting the epigenetic silencing machinery to promote pluripotency. : Zhang et al. report an interplay between the DEAD-box helicase DDX18 and the polycomb repressive complex 2 (PRC2) in governing the hyperactive state of ribosomal DNA in pluripotency regulation. DDX18 counteracts PRC2 complex formation and prevents PRC2 binding and aberrant histone methylation at rDNA loci. Keywords: stem cell pluripotency, DDX18, RNA-binding protein, PRC2, rRNA transcription, rDNA loci, nucleolus

Transcription regulation underlies stem cell function and development. Here, we elucidate an unexpected role of an essential ribogenesis factor, WDR43, as a chromatin-associated RNA-binding protein (RBP) and release factor in modulating the polymerase (Pol) II activity for pluripotency regulation. WDR43 binds prominently to promoter-associated noncoding/nascent RNAs, occupies thousands of gene promoters and enhancers, and interacts with the Pol II machinery in embryonic stem cells (ESCs). Nascent transcripts and transcription recruit WDR43 to active promoters, where WDR43 facilitates releases of the elongation factor P-TEFb and paused Pol II. Knockdown of WDR43 causes genome-wide defects in Pol II release and pluripotency-associated gene expression. Importantly, auxin-mediated rapid degradation of WDR43 drastically reduces Pol II activity, precluding indirect consequences. These results reveal an RNA-mediated recruitment and feedforward regulation on transcription and demonstrate an unforeseen role of an RBP in promoting Pol II elongation and coordinating high-level transcription and translation in ESC pluripotency.

SummarySuper-enhancers (SEs) are large clusters of transcriptional enhancers that are co-occupied by multiple lineage-specific transcription factors driving expression of genes that define cell identity. In embryonic stem cells (ESCs), SEs are highly enriched for the core pluripotency factors Oct4, Sox2, and Nanog. In this study, we sought to dissect the molecular control mechanism of SE activity in pluripotency and reprogramming. Starting from a protein interaction network surrounding Sox2, we identified Tex10 as a key pluripotency factor that plays a functionally significant role in ESC self-renewal, early embryo development, and reprogramming. Tex10 is enriched at SEs in a Sox2-dependent manner and coordinates histone acetylation and DNA demethylation at SEs. Tex10 activity is also important for pluripotency and reprogramming in human cells. Our study therefore highlights Tex10 as a core component of the pluripotency network and sheds light on its role in epigenetic control of SE activity for cell fate determination.

Pervasive transcription in mammalian genomes produces thousands of long noncoding RNA (lncRNA) transcripts. Although they have been implicated in diverse biological processes, the functional relevance of most lncRNAs remains unknown. Recent studies reveal the prevalence of lncRNA-mediated cis regulation on nearby transcription. In this review, we summarize cis- and trans-acting lncRNAs involved in stem cell pluripotency and reprogramming, highlighting the role of regulatory lncRNAs in providing an additional layer of complexity to the regulation of genes that govern cell fate during development.

Ten-eleven translocation (TET) proteins play key roles in the regulation of DNA-methylation status by oxidizing 5-methylcytosine (5mC) to generate 5-hydroxymethylcytosine (5hmC), which can both serve as a stable epigenetic mark and participate in active demethylation. Unlike the other members of the TET family, TET2 does not contain a DNA-binding domain, and it remains unclear how it is recruited to chromatin. Here we show that TET2 is recruited by the RNA-binding protein Paraspeckle component 1 (PSPC1) through transcriptionally active loci, including endogenous retroviruses (ERVs) whose long terminal repeats (LTRs) have been co-opted by mammalian genomes as stage- and tissue-specific transcriptional regulatory modules. We found that PSPC1 and TET2 contribute to ERVL and ERVL-associated gene regulation by both transcriptional repression via histone deacetylases and post-transcriptional destabilization of RNAs through 5hmC modification. Our findings provide evidence for a functional role of transcriptionally active ERVs as specific docking sites for RNA epigenetic modulation and gene regulation.

To analyze the association between serum levels of S100A8/S100A9 and clinicopathological features of colorectal cancer patients. A total of 82 patients with CRC and 14 healthy controls were enrolled for this study. The levels of S100A8 and S100A9 in serum were detected by ELISA assay. The association between S100A8/S100A9 and clinicopathological features was analyzed by student-t test and one-way ANOVA. Receiver Operating Characteristic curve was used to analyze diagnostic efficiency of serum S100A8 and S100A9 for colon rectal cancer. Logistic regression model was also established to analyze the possible risk factors for elevation of S100A8/S100A9. The levels of S100A8 and S100A9 were (1 403.3±593.7) and (2 890.3±994.9) pg/mL in patients with colon cancer, and (712.8±265.3) and (1 492.7±564.6) pg/mL in controls, respectively, with significant difference between the two groups (P0.01). The similar results were found in rectal cancer patients, with a level of S100A8 and S100A9 at (1 417.7±666.5) and (3 026.7±887.6) pg/mL, respectively. Diagnostic sensitivity and specificity of S100A8 and S100A9 are better than traditional biomarkers. The levels of S100A9 in serum of CRC patients were correlated with clinical stages and distant metastasis. Serum levels of S100A9 in patients of stage III [(3 111.9±178.5) pg/mL] and stage IV [(3 831.4±278.5) pg/mL] were significantly (P0.01) higher than that in stage I [(2 276.1±167.4) pg/mL], whereas there was significant change in S100A8 levels. Logistic regression showed the possible risk factors for the elevation of S100A9, including depth of invasion, lymphatic metastasis and degree of differentiation (P0.05). Serum level of S100A8 and S100A9 in CRC patients were significantly increased and serum level of S100A9 was positively correlated with the malignant features of CRC.目的：探讨血清S100A8和S100A9表达水平和结直肠癌患者临床特征的关系。方法：用ELISA法检测82例结肠癌或直肠癌患者和14例健康对照者血清中S100A8和S100A9的含量，利用t检验、方差分析等方法分析两种分子与结直肠癌病理分期及临床特征的关系；利用受试者工作特征曲线(ROC曲线)分析S100A8和S100A9对结肠癌和直肠癌的诊断效能；构建logistic回归模型分析S100A8和S100A9血清水平升高的可能相关因素。结果：结肠癌组患者血清中S100A8浓度[(1 403.3±593.7) pg/mL]和S100A9的含量[(2 890.3±994.9) pg/mL]均显著高于健康对照组[分别为(712.8±265.3)和(1 492.7±564.6) pg/mL，P0.01]；直肠癌患者血清S100A8和S100A9的含量分别是(1 417.7±666.5)和 (3 026.7±887.6) pg/mL，也高于对照组(P0.05)；血清S100A8和S100A9诊断结肠癌或直肠癌均具有较高的敏感性和特异性，两者联合应用时，曲线下面积(AUC)在结肠癌和直肠癌分别达到0.972和0.964；随着结直肠癌临床恶性度增高和远处转移的发生，血清S100A9表达水平升高，差异具有统计学意义(P0.05)，S100A9在III期[(3 111.9±178.5) pg/mL]和IV期[(3 831.4±278.5) pg/mL]患者血清中的含量均显著高于I期患者[(2 276.1±167.4) pg/mL，P0.01]；而S100A8血清浓度与临床分期之间未发现明显关联；logistic回归分析发现血清S100A9升高与肿瘤浸润深度(OR=6.135，P=0.024)、淋巴结转移(OR=4.735，P=0.05)及分化程度(OR=2.573，P=0.043)等密切相关。结论：结直肠癌患者的S100A8和S100A9血清水平显著升高，且S100A9水平与结直肠癌恶性度呈正相关。.

Summary Endothelial progenitor cells (EPCs) play a protective role in the cardiovascular system by enhancing the maintenance of endothelium homeostasis and the process of new vessel formation. Recent studies show that EPCs may induce vascular regeneration and neovascularization mainly through paracrine signaling, that is, through the secretion of growth factors and pro-angiogenic cytokines [1]. However, multiple factors might function synergistically and therefore make it difficult to manipulate EPC paracrine effects. MicroRNAs, a family of small, non-coding RNAs, are characterized by post-transcriptionally regulating multiple functionally related genes, which renders them potentially powerful therapeutic targets or tools. In this paper we propose the hypothesis that microRNAs can be utilized as a novel therapeutic strategy for regulating EPC paracrine secretion.

The unfolded protein response (UPR) is an intracellular stress-signaling pathway that counteracts the accumulation of misfolded proteins in the endoplasmic reticulum (ER). Because defects in ER protein folding are associated with many pathological states, including metabolic, neurologic, genetic, and inflammatory diseases, it is important to understand how the UPR maintains ER protein-folding homeostasis. All metazoans have conserved the fundamental UPR transducers IRE1, ATF6, and PERK. In Caenorhabditis elegans , the UPR is required to prevent larval lethality and intestinal degeneration. Although ire-1 -null worms are viable, they are particularly sensitive to ER stress. To identify genes that are required for development of ire-1 -null worms, we performed a comprehensive RNA interference screen to find 10 genes that exhibit synthetic growth and intestinal defects with the ire-1(v33) mutant but not with atf-6(tm1153) or pek-1(ok275) mutants. The expression of two of these genes, exos-3 and F48E8.6 , was induced by ER stress, and their knockdown in a wild-type strain caused ER stress. Because these genes encode subunits of the exosome complex that functions in mRNA surveillance, we analyzed other gene products required for nonsense-mediated mRNA decay (NMD). Our results demonstrate that defects in smg-1 , smg-4 , and smg-6 in C. elegans and SMG6 in mammalian cells cause ER stress and sensitize to the lethal effects of ER stress. Although ER stress did not activate mRNA surveillance complex assembly, ER stress did induce SMG6 expression, and NMD regulators were constitutively localized to the ER. Importantly, the findings demonstrate a unique and fundamental interaction where NMD-mediated mRNA quality control is required to prevent ER stress.

SummaryEpigenetic alterations have been increasingly implicated in oncogenesis. Analysis of Drosophila mutants suggests that Polycomb and SWI/SNF complexes can serve antagonistic developmental roles. However, the relevance of this relationship to human disease is unclear. Here, we have investigated functional relationships between these epigenetic regulators in oncogenic transformation. Mechanistically, we show that loss of the SNF5 tumor suppressor leads to elevated expression of the Polycomb gene EZH2 and that Polycomb targets are broadly H3K27-trimethylated and repressed in SNF5-deficient fibroblasts and cancers. Further, we show antagonism between SNF5 and EZH2 in the regulation of stem cell-associated programs and that Snf5 loss activates those programs. Finally, using conditional mouse models, we show that inactivation of Ezh2 blocks tumor formation driven by Snf5 loss.

Trimethylation on histone H3 lysine 27 (H3K27me3) by Polycomb repressive complex 2 (PRC2) regulates the balance between self-renewal and differentiation of embryonic stem cells (ESCs). The mechanisms by which the activity and recruitment of PRC2 are controlled are largely unknown. Here we demonstrate that the founding member of the Jumonji-family, JMJ (JUMONJI or JARID2), is tightly associated with PRC2, colocalizes with PRC2 and H3K27me3 on chromatin, and modulates PRC2 function. In vitro JMJ inhibits PRC2 methyltransferase activity, a finding consistent with increased H3K27me3 marks at PRC2 targets in Jmj−/− ESCs. Paradoxically, JMJ is required for efficient binding of PRC2, indicating that the interplay of PRC2 and JMJ fine-tunes deposition of the H3K27me3 mark. During differentiation, activation of genes marked by H3K27me3 and lineage commitments are delayed in Jmj−/− ESCs. Our results demonstrate that dynamic regulation of Polycomb complex activity orchestrated by JMJ balances self-renewal and differentiation, further highlighting the involvement of chromatin dynamics in cell-fate transitions.

Objective: To evaluate the effects of community-based rehabilitation therapy on neurological function deficit in stroke patients. Design: Prospective, single-blind, randomized controlled multicentre trial. Setting: At home, in Shanghai, China. Subjects: A total of 737 stroke patients in the community. Intervention: The rehabilitation group received additional standardized community-based rehabilitation therapy at home for five months. Main outcome measures: Patients were evaluated using the Clinical Neurological Function Deficit Scale before intervention and at the end of two and five months. Results: Although both the rehabilitation group and the control group improved over time, the rehabilitation group showed a greater improvement in Clinical Neurological Function Deficit Scale scores. The differences between the groups were significant. After five months, the Clinical Neurological Function Deficit Scale scores of the cerebral infarction rehabilitation group improved by 6.77; the haemorrhage rehabilitation group by 7.99; the total rehabilitation group by 7.03. In comparison, the Clinical Neurological Function Deficit Scale scores of the cerebral infarction control group improved by 1.57; the haemorrhage control group by 5.34; the total control group by 2.43. This implies a difference in improvement of 5.2 in the cerebral infarction group, 2.65 in the haemorrhage group, and 4.6 in the total group in favour of the rehabilitation group between groups. Conclusion: Standardized community-based rehabilitation therapy may help stroke patients to improve their neurological function.

Increasing evidence suggests endothelial progenitor cells (EPCs) improve neovascularization and endothelium regeneration. Resveratrol (RSV) is a natural polyphenolic compound, which has been demonstrated to exert multiple protective effects on the cardiovascular system, including inhibition of platelet adhesion and aggregation, reduction of myocardial ischemia‑reperfusion injury, and suppression of neointimal hyperplasia of injured vascular tissue. The present study investigated the role of RSV on levels of oxidative stress and senescence of EPCs, and the effects of RSV on vascular‑promoting and/or vascular‑healing capacity of EPCs. It was demonstrated that EPCs could promote the repair of endothelium of the injured artery. RSV reduced the oxidative reaction of EPCs and inhibited EPC senescence, and these effects may occur via the peroxisome proliferator‑activated receptor‑γ/heme oxygenase‑1 signaling pathways.

The endoplasmic reticulum (ER) is a factory for folding and maturation of newly synthesized transmembrane and secretory proteins. The ER provides stringent quality control systems to ensure that only correctly folded proteins exit the ER and unfolded or misfolded proteins are retained and ultimately degraded. A number of biochemical and physiological stimuli can change ER homeostasis, impose stress to the ER, and subsequently lead to accumulation of unfolded or misfolded proteins in the ER lumen. The ER has evolved stress response signaling pathways collectively called the unfolded protein response (UPR) to cope with the accumulation of unfolded or misfolded proteins. This review summarizes our understanding of the UPR signaling developed in the recent years.

Recent studies have suggested that endothelial progenitor subpopulation (EPCs) number and activity were associated with EPCs senescence. Our previous study had shown that stromal cell-derived factor-1alpha (SDF-1α) could prevent EPCs senescence, which may be via telomerase. In this study, we further investigated the role of human telomerase reverse transcriptase (h-TERT) on the protective effect of SDF-1α against senescence. Knockdown h-TERT abrogated the protective effect of SDF-1α and abolished the effects of SDF-1α on migration and proliferation. Moreover, it inhibited EPCs recruitment. In conclusion, h-TERT served a critical role in the progress that SDF-1α prevented EPCs senescence and enhanced re-endothelialization of the injured arteries.

Objective: We retrospectively analyzed the surgical outcome and safety of the Montgomery T-tube for laryngotracheal stenosis. Methods: The patients with laryngotracheal stenosis who had undergone T-tube placement between 1996 and 2010 were reviewed. The severity of the stenosis was evaluated using the Cotton-Myer staging method. The primary endpoint was the rate of successful extubation and the secondary endpoint was safety. Results: 546 patients were eligible. T-tubes were successfully extubated in 342 patients 6-24 months following intubation. The initial extubation success rate was 62.3%. Laryngotracheal restenosis following extubation occurred in 192 patients, necessitating T-tube placement for a second time. The success rate for the second attempt was 58.9%. The overall success rate was 83.3%. Hemoptysis was reported in 8 patients, postoperative infection in 6 patients, wound dehiscence in 3 patients, laryngeal obstruction in 13 patients, aspiration in 12 patients, and postoperative tracheoesophageal fistula in 2 patients. Conclusion: This large clinical series demonstrated the safety and effectiveness of the T-tube for grade 1 and 2 stenosis with stenosed segments of 6 cm, tracheal end-to-end anastomosis is not appropriate and long-term placement of a T-tube is recommended. Our findings provide useful guidance for preoperatively selecting patients with laryngotracheal stenosis of various causes and differing severity.

Histone H3 lysine 27 (H3K27) methylation and H2A monoubiquitination (ubH2A) are two closely related histone modifications that regulate Polycomb silencing. Previous studies reported that H3K27 trimethylation (H3K27me3) rarely coexists with H3K36 di- or tri-methylation (H3K36me2/3) on the same histone H3 tails, which is partially controlled by the direct inhibition of the enzymatic activity of H3K27-specific methyltransferase PRC2. By contrast, H3K27 methylation does not affect the catalytic activity of H3K36-specific methyltransferases, suggesting other Polycomb mechanism(s) may negatively regulate the H3K36-specific methyltransferase(s). In this study, we established a simple protocol to purify milligram quantities of ubH2A from mammalian cells, which were used to reconstitute nucleosome substrates with fully ubiquitinated H2A. A number of histone methyltransferases were then tested on these nucleosome substrates. Notably, all of the H3K36-specific methyltransferases, including ASH1L, HYPB, NSD1, and NSD2 were inhibited by ubH2A, whereas the other histone methyltransferases, including PRC2, G9a, and Pr-Set7 were not affected by ubH2A. Together with previous reports, these findings collectively explain the mutual repulsion of H3K36me2/3 and Polycomb modifications.

Senescence is an irreversible growth arrest which can be triggered by stresses such as oxidative reaction, telomere shortening, DNA damage, or oncogene signaling. Oxidative stress accelerates vascular endothelial cell senescence, and may promote atherosclerosis in humans. Interleukin-8 (IL-8) has been shown to play an important role in tumor growth, angiogenesis, and metastasis, and has close relationship with oxidative stress. The objective of this study was to determine if IL-8 might be able to prevent oxidative stress-induced senescence of endothelial cells and the mechanisms. Human umbilical vein endothelial cells (HUVECs) were cultured and stimulated with hydrogen peroxide in the absence or presence of IL-8. After ex vivo cultivation, HUVECs became senescent as determined by acidic beta-galactosidase staining. IL-8 dose-dependently inhibited the onset of HUVEC senescence. Western blots indicated that IL-8 attenuated the oxidative stress induced high-expression of cell cycle regulation protein and inhibited the activation of p38 and NF-κB pathway. IL-8 also increased telomerase activity which was accompanied with upregulation of the catalytic subunit, telomerase reverse transcriptase (TERT), whereas these effects were significantly attenuated by SB 225002 (selective non-peptide CXCR2 antagonist). In conclusion, IL-8 exerted protective effects against endothelial senescence, which may be related to the activation of telomerase.

The discovery of endothelial progenitor cells (EPCs) provides us with a novel treatment strategy for complications requiring therapeutic revascularization and vascular repair. However, the feasibility of this strategy may be limited due to reduced number and impaired function of EPCs under stimulation of TNF-α. The present study was designed to investigate the effect of rosiglitazone on EPC apoptosis induced by TNF-α and the molecular mechanisms involved. Rosiglitazone attenuated apoptosis of TNF-α–stimulated EPCs in a dose-dependent manner. Rosiglitazone decreased caspase-3 activity and cleavages of caspase-3, caspase-7, and parp. Rosiglitazone also moderated the dissipation of mitochondrial membrane potential caused by TNF-α treatment and reduced the expression of bax and the release of cytochrome c. Furthermore, rosiglitazone inhibited phosphorylations of ERK/MAPK and NF-κB signal molecules. Both ERK and NF-κB inhibitors decreased TNF-α–induced apoptosis of EPCs, as well as the expression of cleaved caspase-3 and parp. These results suggest that rosiglitazone may mediate the inhibitory effect on EPCs apoptosis under TNF-α stimulation through suppression of ERK/MAPK and NF-κB signal pathways. Keywords:: endothelial progenitor cell, TNF-α, rosiglitazone, apoptosis, molecular mechanism

To study how to prevent recurrent laryngeal nerve (RLN) lesions by anastomosing it during thyroid surgery.In the present study 517 patients with thyroid diseases underwent thyroid surgery from January 1993 to May 2005, with RLNs of 163 cases (187 sides, A group) anatomized and RLNs of 354 cases (438 sides. B group) not anatomized. The RLN in B group were protected generally during thyroid surgery. RLNs of A group were anatomized partly or totally.(1) In A group RLNs of 123 cases were partly anatomized and 64 totally. None of all RLNs was injuried. (2) B group 3 sides of 3 cases of all RLNs was injuried. The rate of RLN's lesion is 0.7%, which is significantly higher in A group than in B group.Anastomosing RLN during thyroid surgery may prevent its lesions. The length of anatomized RLN must vary with the area and position of thyroid pathological changes. The RLN needn't be anatomized in these patients with the benign thyroid pathological changes which is far away trachea-oesophagus channel.

Embryonic stem (ES) cells are pluripotent and of therapeutic potential in regenerative medicine. Understanding pluripotency at the molecular level should illuminate fundamental properties of stem cells and the process of cellular reprogramming. Through cell fusion the embryonic cell phenotype can be imposed on somatic cells, a process promoted by the homeodomain protein Nanog, which is central to the maintenance of ES cell pluripotency. Nanog is thought to function in concert with other factors such as Oct4 (ref. 8) and Sox2 (ref. 9) to establish ES cell identity. Here we explore the protein network in which Nanog operates in mouse ES cells. Using affinity purification of Nanog under native conditions followed by mass spectrometry, we have identified physically associated proteins. In an iterative fashion we also identified partners of several Nanog-associated proteins (including Oct4), validated the functional relevance of selected newly identified components and constructed a protein interaction network. The network is highly enriched for nuclear factors that are individually critical for maintenance of the ES cell state and co-regulated on differentiation. The network is linked to multiple co-repressor pathways and is composed of numerous proteins whose encoding genes are putative direct transcriptional targets of its members. This tight protein network seems to function as a cellular module dedicated to pluripotency.

The clinical effects of three different approaches were retrospectively analysed in the treatment of non-invasive paranasal sinuses mycosis.Thirty-two cases between January, 1999 to December, 2003 with non-invasive paranasal sinuses were operated by using nasal endoscope in different approaches, such as two pathway of in-nose and out-nose,two pathway of middle and inferior meatus, and single pathway of middle meatus. The clinical effects were analysed.The three different approaches were adopted according to different nasal cavity structure and good effects were got.No matter which operation styles is adopted, the final aim is to drainage and to ventilate the sinuses, to clean the pathological lesion and to make the wound as slight as possible.

To analyze the role of chemokines and their receptors in the mechanism of allergic rhinitis (AR) by observing the expression of genes of chemokines and their receptors in nasal mucosa of AR through gene chip.The total RNAs were isolated from nasal mucosa of AR and purified to mRNAs, then reversely transcribed to cDNAs and incorporated with fluorescent-labled CY5-dUPT for probes preparion. CY3-dUTP probes prepared with normal nasal mucosa of normal for control. The chip contains cDNAs of chemokines and their receptors were used to hybridized with probes then screened with computer to study the expression of genes based on different density of fluorescent.The chemokines of CCL11 (eotaxin-1), CCL24 (eotaxin-2), CCL7 (MCP-3), CCL13 (MCP-4), RANTES (CCL5) and receptors of CCR2, CCR3, CCR4, CCR5 were differential expressed on four samples, and most of the chemokines and their receptors has tendency regulation on T help 2 (Th2) lymphocytes and involved in the prodeeds such as inducement of allergic reaction,accumulation of inflammacytes and degranulation of sensitized cells.A disorder exists in T helper immune system with AR. The chemokines and their receptors that polarized with Th2 lymphocytes perhaps play important roles in AR pathogenesis, and it represents a new approach to AR immunotherapy.

To study the clinical manifestation, diagnosis and treatment of laryngeal neuroendocrine neoplasms.Three cases of laryngeal neuroendocrine neoplasms were analyzed, of which two cases were diagnosed as atypical carcinoids with radical surgery and neck dissection, another case was diagnosed as asparaganglioma through local neoplastic removal.Two patients of atypical carcinoids are in a good condition after following up one year and another patient of asparaganglioma is with good prognosis after following up eleven years.Laryngeal neuroendocrine neoplasms are composed of a morphologically heterogeneous group of lesions. Accurate diagnosis is primarily based on clinical characteristic, microscopic and immunohistochemical examination. In some instances, it may be supported by ultrastructural studies. Difference treatment should be provided to different classified tumor.

To explore the effect of Diomed-25 semi-conductive laser in treatment of vocal cord leukoplakia.From 1997 to 2001, 30 patients with vocal cord leukoplakia were operated in our hospital by Diomed-25 laser, acoustic analysis were compared and studied before and after operation. All the patients were followed up, ranging from 1 to 5 years.There were 16 cases of simple leukoplakia and 14 cases with mild, moderate or severe atypical hyperplasia in this study. None of simple leukoplakia and mild atypical hyperplasia had malignant change, but 2 cases had malignant change who were the cases with moderate and severe atypical hyperplasia respectively. Acoustic analysis showed significant difference between preoperation and postoperation group (P0.05).The Diomed-25 laser was proved valuable in treating patients with vocal cord leukoplakia.

To investigate 5 procedures of laryngeal reinnervation for unilateral vocal cord paralysis induced by traumatic recurrent laryngeal nerve injury.35 cases were selected for our study, all patients had unilateral recurrent laryngeal nerve injury, including 8 for nerve decompression, 6 for end to end anastomosis of recurrent laryngeal nerve, 16 for main branch of ansa cervicalis anastomosis to recurrent laryngeal nerve, 3 for nerve muscular pedicle and 2 for nerve implantation. All cases have been subjected to preoperative and postoperative voice recording, acoustic analysis, videolaryngoscopy, strobscopy and electromyography.It is found the adductory and abductory motion of the vocal cord restored in 5 cases with less than 4 months course who received nerve decompression. Although functional motion of vocal cord was not seen in two patients who received nerve decompression with a course longer than 4 months and one less than 4 months, and in all cases who received ansa cervicalis anastomosis and end to end anastomosis of recurrent laryngeal nerve, these procedures resulted in medialization of vocal cord and the mass and tension of the reinnervated vocal cord may become much the same as the contralateral normal vocal cord, thus resuming symmetric vibration of the vocal cords and physiological phonation. Nerve muscular pedicle technique and nerve implantation enabled adductory muscles to be reinnervated, thus improving severe hoarseness, but they didn't restore normal voice.(1) Nerve decompression seems to be the best procedure in laryngeal reinnervation; (2) Main branch of ansa cervicalis technique raises satisfactory reinnervation of adductor muscles; (3) Selection of the laryngeal reinnervation protocols should depend on the course, severity and type of nerve injury.

To reestablish the respiratory abduction of the paralyzed vocal cord through reinnervation of the posterior cricoarytenoid(PCA) muscle by the phrenic nerve in humans.In six cases with bilateral recurrent laryngeal nerve paralysis, the phrenic nerve was anastomosed to the anterior branch of recurrent laryngeal nerve, while the adductor branch of recurrent laryngeal nerve was severed and its proximal end was implanted into the PCA muscle belly in one side, for the other side nerve-muscle pedicle technique was used. All cases had been subjected to preoperative and postoperative video laryngoscopy, stroboscopy, electromyography, voice recording and acoustic analysis.Among the 6 patients, it is observed in five cases' phrenic nerve reinnervation side the inspiratory abducent motion evidently recovered, and the abducent range was from 3 to 5 mm, While only slight abductent motion or no motion could be recorded on the other side reinnervated with nerve-muscle pedicle technique, and the vocal cord excursion on this side was less than 1 mm in all cases. It is because the glottis is broad enough for the patients to have daily activities without short of breath, so all of them were decannulated postoperatively. The reinnervated PCA muscle by the phrenic nerve showed typical inspiratory high frequency discharge with 100-200 ms delay as compared with the other side, indicating the phrenic motoneuron pattern. No long-term diaphragmatic paralysis and lesion of respiratory function was found. All cases' voice was not weakened, and no aspiration occurred.The phrenic reinnervation is feasible clinically for treating vocal cord paralysis, and it is found to be more effective for restoring inspiratory abducent function than the nerve-muscle pedicle technique.

To discover the etiology of allergic rhinitis(AR) at the genomic level.Microarray analysis on a genomic scale was used to find changes in genes expression in AR. The total RNAs were isolated from nasal mucosa of AR and normal, then purified to mRNAs and were reversely transcribed to cDNAs with the incorporation of fluorescent-labeled dUTP to prepare for the hybridization probes. The genes differential expressed in AR were screened with the gene chip which contained 12,800 human genes in all 8 samples.734 genes were shown in differential expressed profile with 430 upregulated genes and 304 downregulated genes, and all 8 samples were 15 genes upregulated and 6 genes downregulated. The differential expressed genes involved in the functions of inflammation, immunology, neuroendocrine and signal conduct, and they had potential value in AR study.The investigation based on gene chip in researching related genes in AR afforded a new idea in studying pathogenesis of nasal allergic diseases.

To explore the protocols and effects of laryngeal reinnervation for unilateral recurrent laryngeal nerve (RLN) injury caused by thyroid surgery.Different protocols of laryngeal reinnervation were performed upon 29 patients with unilateral recurrent laryngeal nerve injury caused by thyroid surgery, just coming on to with a course of 2 years, including nerve decompression upon 8 cases, end to end anastomosis of recurrent laryngeal nerve upon 6 cases, and anastomosis of main branch of ansa cervicalis to recurrent laryngeal nerve upon 15 cases. All were been subjected to preoperative and postoperative voice recording, acoustic analysis, videolaryngoscopy, stroboscopy and electromyography.Nerve decompression had restored the normal functional adductory and abductory motion of the vocal cord in 5 patients with a course of less than four months. Although functional motion of vocal cord had not been recovered in three patients who received nerve decompression, 2 being with a course of longer than 4 months and one with a course of less than 4 months, and in all cases who received ansa cervicalis anastomosis and end to end anastomosis of recurrent laryngeal nerve, these procedures resulted in medialization of vocal cords except in two cases, one receiving ansa cervicalis anastomosis, and the other receiving end to end anastomosis of RLN. Acoustic parameters (Jitter, Shimmer, NNE) measured 6 months after operation of any kind all returned to normal, however, without significant difference among different groups (P0.05). The amplitude of evoked potential of reinnervated laryngeal muscles was significantly greater in the group of nerve decompression than in the group of end to end anastomosis of RLN and group of ansa cervicalis anastomosis (both P0.05). However, the amplitude of evoked potential of reinnervated laryngeal muscles between the latter two groups was not significantly different (P0.05). Except in one case in the end to end anastomosis of RLN group and one case in the ansa cervicalis anastomosis group, the mass and tension of the reinnervated vocal cord became much the same as the contralateral normal vocal cord and symmetric vibration of the vocal cords and physiological phonation were recovered in all cases.(1) Nerve decompression is the best procedure in laryngeal reinnervation. (2) Main branch of ansa cervicalis anastomosis and end to end anastomosis of RLN effectively restore the laryngeal vocalization. (3) Selection of the laryngeal reinnervation protocols should depend on the course, severity and type of nerve injury.

Histone modifications have been implicated in stem cell maintenance and differentiation. We have analyzed genome-wide changes in gene expression and histone modifications during differentiation of multipotent human primary hematopoietic stem/progenitor cells (HSCs/HPCs) into erythrocyte precursors. Our data indicate that H3K4me1, H3K9me1 and H3K27me1 associate with enhancers of differentiation genes prior to their activation and correlate with basal expression, suggesting that these mono-methylations are involved in the maintenance of activation potential required for differentiation. In addition, although the majority of genes associated with both H3K4me3 and H3K27me3 in HSCs/HPCs become silent and lose H3K4me3 after differentiation, those that lose H3K27me3 and become activated after differentiation are associated with increased levels of H2A.Z, H3K4me1, H3K9me1, H4K20me1 and RNA polymerase II in HSCs/HPCs. Thus, our results suggest that gene expression changes during differentiation are programmed by chromatin modifications present at the HSC/HPC stage and we provide a resource for enhancer and promoter identification.

Recent studies have shown that patients whose bladder cancer exhibit overexpression of RB protein as measured by immunohistochemical analysis do equally poorly as those with loss of RB function. We hypothesized that loss of p16 protein function could be related to RB overexpression, since p16 can induce transcriptional downregulation of RB and its loss may lead to aberrant RB regulation. Conversely, loss of RB function has been associated with high p16 protein expression in several other tumor types. In the present study RB negative bladder tumors also exhibited strong nuclear p16 staining while each tumor with strong, homogeneous RB nuclear staining were p16 negative, supporting our hypothesis. To expand on these immunohistochemical studies additional cases were selected in which the status of the p16 encoding gene had been determined at the molecular level. Absent p16 and high RB protein expression was found in the tumors having loss of heterozygosity within 9p21 and a structural change (mutation or deletion) of the remaining p16 encoding gene allele, confirming the staining results. These results strongly support the hypothesis that the RB nuclear overexpression recently associated with poor prognosis in bladder cancer is also associated with loss of p16 function and implies that loss of p16 function could be equally deleterious as RB loss in bladder and likely other cancers.