Timing of MeCP2 Expression Determines Midbrain Dopamine Neuron Phenotype Specification.

Midbrain dopamine (DA) neurons are associated with locomotor and psychiatric disorders. DA phenotype is specified in ancestral neural precursor cells (NPCs) and maintained throughout neuronal differentiation. Here we show that endogenous expression of MeCP2 coincides with DA phenotype specification in mouse mesencephalon, and premature expression of MeCP2 prevents in vitro cultured NPCs from acquiring DA phenotype through interfering NURR1 transactivation of DA phenotype genes. By contrast, ectopic MeCP2 expression does not disturb DA phenotype in the DA neurons. By analyzing the dynamic change of DNA methylation along DA neuronal differentiation at the promoter of DA phenotype gene tyrosine hydroxylase (Th), we show that Th expression is determined by TET1-mediated de-methylation of NURR1 binding sites within Th promoter. Chromatin immunoprecipitation assays demonstrate that premature MeCP2 dominates the DNA binding of the corresponding sites thereby blocking TET1 function in DA NPCs, whereas TET1-mediated de-methylation prevents excessive MeCP2 binding in DA neurons. The significance of temporal DNA methylation status is further confirmed by targeted methylation/demethylation experiments showing that targeted de-methylation in DA NPCs protects DA phenotype specification from ectopic MeCP2 expression, whereas targeted methylation disturbs phenotype maintenance in MeCP2-overexpressed DA neurons. These findings suggest the appropriate timing of MeCP2 expression as a novel determining factor for guiding NPCs into DA lineage.

A novel histone deacetylase inhibitor-based approach to eliminate microglia and retain astrocyte properties in glial cell culture.

The close association between astrocytes and microglia causes great difficulties to distinguish their individual roles in innate immune responses in central nervous system. Current chemical-based methods to eliminate microglia in glial cell culture introduce various molecular and functional alterations to astrocytes. Here, we describe a novel two-step approach to achieve a complete elimination of microglia without affecting the biological properties of co-cultured astrocytes by temporal treatment of histone deacetylase inhibitor trichostatin A (TSA). We verify TSA as a potent inducer for microglial-specific apoptotic cell death, which also causes comprehensive gene expression changes in astrocytes. However, withdrawal of TSA not only ensures no microglia repopulation, but also restores all the gene expression changes in terms of astrocyte functions, including neurotrophic factors, glutamate and potassium transporters, and reactive astrocyte subtypes. By contrast, withdrawal of PLX5622, the commonly used colony-stimulating factor 1 receptor inhibitor neither prevents microglia repopulation nor restores the gene expression changes mentioned above. Using this method, we are able to discriminate differential roles of microglia and astrocytes in the induced expression of antiviral and pro-inflammatory cytokines upon various pathological stimuli including the spike protein of SARS-CoV-2. This simple and efficient method can be customized for the understanding of microglia-astrocyte interaction and the development of epigenetic therapies that target over-activated microglia in neuroinflammation-related diseases.

Vitamin C Promotes Astrocyte Differentiation Through DNA Hydroxymethylation.

Previous studies have reported that vitamin C (VC) promotes neural stem/precursor cell (NSC) differentiation toward dopamine (DA) neurons via DNA hydroxymethylation-induced transcriptional activation of DA neuron-specific genes. To further understand the VC effects on NSC differentiation, we profiled the transcriptome and DNA methylome/hydroxymethylome using high-throughput sequencing. Interestingly, RNA sequencing analyses have shown that, in addition to DA neuronal genes, astrocytic genes Gfap, Slc1a3, and S100a16 were also upregulated in NSC cultures differentiated with VC treatment. Consistently, enhanced GFAP+ astrocytic yields were manifested in the differentiated cultures with VC treatment, collectively indicating that VC promotes astrocytic differentiation. In genome-wide hydroxymethylome analyses, VC treatment induces enrichment of DNA hydroxymethylation (5-hydroxymethyl cytosine; 5hmC) near the consensus binding motifs of nuclear factor I (NFI). Furthermore, we showed that VC significantly enhanced recruitment of NFI and STAT3, key transcription factors for astrogenesis, in the 5hmC-enriched regions of the astrocyte-specific genes. These findings suggest that VC play important roles in astrocytogenesis during brain development. Stem Cells 2018;36:1578-1588.

Foxa2 acts as a co-activator potentiating expression of the Nurr1-induced DA phenotype via epigenetic regulation.

Understanding how dopamine (DA) phenotypes are acquired in midbrain DA (mDA) neuron development is important for bioassays and cell replacement therapy for mDA neuron-associated disorders. Here, we demonstrate a feed-forward mechanism of mDA neuron development involving Nurr1 and Foxa2. Nurr1 acts as a transcription factor for DA phenotype gene expression. However, Nurr1-mediated DA gene expression was inactivated by forming a protein complex with CoREST, and then recruiting histone deacetylase 1 (Hdac1), an enzyme catalyzing histone deacetylation, to DA gene promoters. Co-expression of Nurr1 and Foxa2 was established in mDA neuron precursor cells by a positive cross-regulatory loop. In the presence of Foxa2, the Nurr1-CoREST interaction was diminished (by competitive formation of the Nurr1-Foxa2 activator complex), and CoREST-Hdac1 proteins were less enriched in DA gene promoters. Consequently, histone 3 acetylation (H3Ac), which is responsible for open chromatin structures, was strikingly increased at DA phenotype gene promoters. These data establish the interplay of Nurr1 and Foxa2 as the crucial determinant for DA phenotype acquisition during mDA neuron development.

SNP rs3202538 in 3'UTR region of ErbB3 regulated by miR-204 and miR-211 promote gastric cancer development in Chinese population.

BACKGROUND/AIMS: ErbB3 is an oncogene which has proliferation and metastasis promotion effects by several signaling pathways. However, the individual expression difference regulated by miRNA was almost still unknown. We focused on the miRNAs associated SNPs in the 3'-UTR of ErbB3 to investigate the further relationship of the SNPs with miRNAs among Chinese gastric cancer (GC) patients. METHODS: We performed case-control study including 851 GC patients and 799 cancer-free controls. Genotyping, real-time PCR assay, cell transfection, the dual luciferase reporter assay, western-blot, cell proliferation and trans-well based cell invasion assay were used to investigate the effects of the SNP on ErbB3 expression. Moreover, a 5-years-overall survival and relapse free survival were investigated between different genotypes. RESULTS: We found that patients suffering from Helicobacter pylori (Hp.) infection indicated to be the susceptible population by comparing with controls. Besides, SNP rs3202538 (G/T) in ErbB3 3'-UTR was involved in the occurrence of GC by acting as tumor risk factors. SNP rs3202538 (G/T) could be regulated by both miR-204 and miR-211 which caused an upregulation of ErbB3 in patients. Furthermore, the carriers of T genotype was related to the significantly high expression of ErbB3, and to big tumor size, poor differentiation as well as the high probability of metastasis. Both miR-211 and miR-204 can significantly decrease cell proliferation, metastasis as well as downstream AKT activation through G but not T allele of ErbB3 3'UTR. Moreover, the SNP of G/T was associated with shorter survival of post-surgery GC patients with 5 years of follow up study. CONCLUSION: In conclusion, our findings have shown that the SNP rs3202538 (G/T) in ErbB3 3'-UTR acted as promotion factors in the GC development through disrupting the regulatory role of miR-204 and miR-211 in ErbB3 expression.

Vitamin C facilitates dopamine neuron differentiation in fetal midbrain through TET1- and JMJD3-dependent epigenetic control manner.

Intracellular Vitamin C (VC) is maintained at high levels in the developing brain by the activity of sodium-dependent VC transporter 2 (Svct2), suggesting specific VC functions in brain development. A role of VC as a cofactor for Fe(II)-2-oxoglutarate-dependent dioxygenases has recently been suggested. We show that VC supplementation in neural stem cell cultures derived from embryonic midbrains greatly enhanced differentiation toward midbrain-type dopamine (mDA) neurons, the neuronal subtype associated with Parkinson's disease. VC induced gain of 5-hydroxymethylcytosine (5hmC) and loss of H3K27m3 in DA phenotype gene promoters, which are catalyzed by Tet1 and Jmjd3, respectively. Consequently, VC enhanced DA phenotype gene transcriptions in the progenitors by Nurr1, a transcription factor critical for mDA neuron development, to be more accessible to the gene promoters. Further mechanism studies including Tet1 and Jmjd3 knockdown/inhibition experiments revealed that both the 5hmC and H3K27m3 changes, specifically in the progenitor cells, are indispensible for the VC-mediated mDA neuron differentiation. We finally show that in Svct2 knockout mouse embryos, mDA neuron formation in the developing midbrain decreased along with the 5hmC/H3k27m3 changes. These findings together indicate an epigenetic role of VC in midbrain DA neuron development.

[Expression of MiR-130a in Serum Samples of Patients with Epithelial Ovarian Cancer and Its Association with Platinum Resistance].

OBJECTIVE: To determine the expression of miR-130a in patients with epithelial ovarian cancer and its association with platinum resistance. METHODS: 32 patients with platinum resistance and 30 patients without platinum resistance were recruited in this study. Real-time PCR was performed to detect the expression of miR-130a in the serum samples of the patients. ELISA was used to measure the expression level of phosphatase and tensin homolog deleted on chromosome ten (PTEN) and B-cell lymphoma-2 (BCL-2). RESULTS: Platinum-resistant patients had significantly higher levels of expression of miR-130a and BCL-2, and lower level of PTEN than platinum-sensitive patients (P < 0.05). The expression level of miR-130a increased with increased severity in histological classification and appearance of lymph node metastasis in the platinum-resistant patients (P < 0.05). CONCLUSION: MiR-130a may mediate the generation of platinum resistance in epithelial ovarian cancer through inhibiting PTEN to activate PI3K/AKT signaling pathway and increasing BCL-2 to inhibit tumor cell apoptosis. MiR-130a may be a new potential target of gene therapy in platinum-resistant ovarian cancers.

Efficacy and safety of probiotics in irritable bowel syndrome: A systematic review and meta-analysis.

BACKGROUND: Irritable bowel syndrome (IBS) is a common gastrointestinal disease characterized by abdominal pain, distension, and altered bowel habits. Probiotics may alleviate IBS symptoms, but clinical trials remain conflicting. AIMS: To conduct a systematic review and meta-analysis of clinical trials to evaluate the efficacy and safety of probiotics for IBS patients. METHODS: We searched relevant trials in PubMed, Web of Science, Embase, Cochrane Library, and Google Scholar from 2000 to June 2023. Standardized mean difference (SMD) and 95% confidence interval (CI) were calculated for continuous outcomes. A risk ratio (RR) and a 95% CI were calculated for dichotomous outcomes. RESULTS: A total of 20 studies involving 3011 patients were obtained. The results demonstrated that probiotics are more effective than placebo in reducing global IBS symptoms improvement rate (RR = 1.401, 95% CI 1.182-1.662, P < 0.001) and quality of life scores (SMD = 0.286, 95% CI = 0.154-0.418, P < 0.001). Subgroup analyses showed that a shorter treatment time (less than eight weeks) could reduce distension scores (SMD = 0.197, 95% CI = 0.038-0.356, P = 0.015). High doses (daily dose of probiotics ≥ 10ˆ10) or multiple strains of probiotics exhibit beneficial effects on abdominal pain (SMD = 0.412, 95% CI = 0.112-0.711, P = 0.007; SMD = 0.590, 95% CI = 0.050-1.129, P = 0.032; respectively). However, there was no significant benefit on global symptom scores (SMD = 0.387, 95% CI 0.122 to 0.653, P = 0.004) with statistically high inter-study heterogeneity (I2 = 91.9%, P < 0.001). Furthermore, there was no significant inter-group difference in terms of adverse events frequency (RR = 0.997, 95% CI 0.845-1.177, P = 0.973). CONCLUSION: Probiotics are effective and safe for IBS patients. High doses or multiple probiotic strains seem preferable, but definite conclusions are challenging due to the high heterogeneity. Large-scale, well-designed, and rigorous trials are needed to confirm their effectiveness.

Prevalence and factors associated with burnout among primary health-care workers in China during COVID-19: a national survey.

BACKGROUND: Primary health-care workers (PHWs) managed increased workloads and pressure during the COVID-19 pandemic. This study conducted a national survey examining burnout among PHWs at the end of the COVID-19 pandemic, and identifies related factors. By doing so, it addresses the gap in understanding the burnout situation among PHWs at a national level, taking into account urban-rural disparities. METHODS: We conducted a nationwide cross-sectional survey of PHWs in China from May to October 2022, covering 31 provinces. The MBI-HSS was used to measure overall burnout and emotional exhaustion (EE), depersonalization (DP), and reduced personal accomplishment (PA). We used multivariable logistic regression to identify risk factors, and subgroup analyses to identify differences between rural and urban areas. RESULTS: 3769 PHWs from 44 primary health-care institutions completed the survey. Overall, 16.6% reported overall burnout, and the prevalence of EE, DP, and reduced PA was 29.7%, 28.0%, and 62.9%, respectively. The prevalence of overall burnout (17.6% vs. 13.7%, P = 0.004) and EE (31.5% vs. 24.8%, P < 0.001) was higher in urban than rural areas (AOR = 1.285; 95%CI, 1.021-1.617). Job satisfaction was a protective factor against burnout in both settings. The protective factors of overall burnout, EE and DP vary between urban and rural areas. CONCLUSIONS: The Mental Health Status Questionnaire-Short Form (MSQ-SF) score functioned as a protective factor against burnout across both rural and urban locales, highlighting the intrinsic link between job satisfaction and burnout. Other influencing factors differed between urban and rural areas, so interventions should be tailored to local conditions. Rural married PHWs experienced the lower prevalence of burnout indicates the support structure may play a significant role. In urban settings, it is recommended to strategically pre-emptively stock essential supplies like PPE.

Discussion on the mechanism of Tiaoqi Xiaowei decoction in the treatment of chronic atrophic gastritis based on network pharmacology and molecular docking: An observational study.

To explore the mechanism of Tiaoqi Xiaowei decoction in the treatment of chronic atrophic gastritis by network pharmacology and molecular docking. The main active components and targets of Tiaoqi Xiaowei decoction were obtained from TCMSP database. The databases of Disgenet, GeneCards, and OMIM were used to obtain chronic atrophic gastritis-related targets. The component-target-disease network was constructed by Cytoscape 3.7.1 software, and the protein-protein interaction network was constructed by String database. The core targets were screened by CytoNCA plug-in. Gene ontology analysis and Kyoto Encyclopedia of Genes and Genome pathway enrichment analysis were performed using the Metascape database. The core components and targets were subjected to molecular docking verification using AutoDock Tools 1.5.6 software, and the binding score was obtained. A total of 48 active components were identified, involving 82 action targets. Core active components such as quercetin, beta-sitosterol, kaempferol, luteolin, and naringenin, and core targets such as AKT1, TP53, VEGFA, TNF, IL6, and PTGS2 were obtained. A total of 188 signaling pathways were screened out, including cancer pathway, PI3K-Akt, IL-17, and TNF signaling pathway. Molecular docking results showed that the key components of Tiaoqi Xiaowei decoction had a favorable binding affinity with key targets. Tiaoqi Xiaowei decoction acts on multiple targets such as AKT1, TP53, VEGFA, TNF, IL6, PTGS2, and synergistically treats chronic atrophic gastritis by regulating inflammatory responses and tumor-related signaling pathways.

[Regulatory effects and associated mechanisms of miR-130a molecules on cisplatin resistance in ovarian cancer A2780 cell lines].

OBJECTIVE: To determine the regulatory effects and associated mechanisms of miR-130a on cisplatin resistance in ovarian cancer A2780 cell lines (including cisplatin sensitive A2780s and its resistant A2780/DDP cells). METHODS: A2780s and A2780/DDP cells were divided into four groups, and treated with lipo2000 (Lip), miR-negative (miR-NC) control, miR-130a-mimics (miR-130a-M increasing the expression of miR-130a and the agent), and miR-130a-inhibitor (miR-130a-I downregulating miR-130a expression), respectively. The proliferation of cells and their sensitivity to cisplatin were detected by MTT assay. RT-PCR and western blot were performed to examine the levels of MDR1, PTEN mRNA and proteins. RESULTS: The expressions of MDR1 mRNA and P-gp in the A2780/DDP cells were significantly higher than those in the A2780s cells. However, no differences in the expressions of PTEN mRNA and proteins were detected between the two cell lines. Over-expressions of miR-130a had no effect on cell proliferation, but increased the resistance of the cells to cisplatin and up-regulated the expressions of MDR1 mRNA and P-gp in both cell lines. Down-regulated miR-130a did not affect cell proliferations, but enhanced the sensitivity of the cells to cisplatin, inhibited the expressions of MDR1 mRNA and P-gp and increased the expression of PTEN proteins. CONCLUSION: MiR-130a expression may be associated with cisplatin resistance of ovarian cancer cells. MiR-130a inhibitor can reverse the cisplatin resistance by upregulating the expression of PTEN proteins and down-regulating P-gp in A2780 cell lines. MiR-130 may become a new potential target of genetic therapy for cisplatin-resistant ovarian cancers.

Prolonged membrane depolarization enhances midbrain dopamine neuron differentiation via epigenetic histone modifications.

Understanding midbrain dopamine (DA) neuron differentiation is of importance, because of physiological and clinical implications of this neuronal subtype. We show that prolonged membrane depolarization induced by KCl treatment promotes DA neuron differentiation from neural precursor cells (NPCs) derived from embryonic ventral midbrain (VM). Interestingly, the depolarization-induced increase of DA neuron yields was not abolished by L-type calcium channel blockers, along with no depolarization-mediated change of intracellular calcium level in the VM-derived NPCs (VM-NPCs), suggesting that the depolarization effect is due to a calcium-independent mechanism. Experiments with labeled DA neuron progenitors indicate that membrane depolarization acts at the differentiation fate determination stage and promotes the expression of DA phenotype genes (tyrosine hydroxylase [TH] and DA transporter [DAT]). Recruitment of Nurr1, a transcription factor crucial for midbrain DA neuron development, to the promoter of TH gene was enhanced by depolarization, along with increases of histone 3 acetylation (H3Ac) and trimethylation of histone3 on lysine 4 (H3K4m3), and decreases of H3K9m3 and H3K27m3 in the consensus Nurr1 binding regions of TH promoter. Depolarization stimuli on differentiating VM-NPCs also induced dissociation of methyl CpG binding protein 2 and related repressor complex molecules (repressor element-1 silencing transcription factor corepressor and histone deacetylase 1) from the CpG sites of TH and DAT promoters. Based on these findings, we suggest that membrane depolarization promotes DA neuron differentiation by opening chromatin structures surrounding DA phenotype genes and inhibiting the binding of corepressors, thus allowing transcriptional activators such as Nurr1 to access DA neuron differentiation gene promoter regions.

[Expression of miR-130a in cisplatin resistant cell lines of ovarian cancer].

OBJECTIVE: To determine the expression of miR-130a in cisplatin resistant cell lines of ovarian cancer and its impact on cisplatin resistance. METHODS: Cisplatin resistant ovarian cancer cell lines were established by stepwise selection with gradual increase of cisplatin. MTT assay was applied to indentify the cisplatin resistant cell lines and determine their resistance index. The expression of miR-130a was measured by SYBR green real-time PCR. RESULTS: The resistance index of A2780/CIS1, A2780/CIS2 and SKOV3/CIS was 30.2, 5.3 and 24.5 respectively. The SYBR green real-time PCR showed that miR-130a was over-expressed in all of the cisplatin resistant cell lines (P < 0.05). The expression of miR-130a was 30.51 times higher in A2780/CIS1, 4.87 times higher in A2780/CIS2 and 24.43 times higher in SKOV3/CIS than in their parental cell lines (P < 0.05), which was almost equally reflected in their resistance index. CONCLUSION: The over expression of miR-130a is associated with cisplatin resistance of ovarian cancer. Inhibiting miR-130a expression may help reverse the cisplatin resistance of ovarian cancer. miR-130a is expected to be a new potential target of genetic therapy for cisplatin resistant ovarian cancer.

Esketamine alleviates postoperative depression-like behavior through anti-inflammatory actions in mouse prefrontal cortex.

The rising incidence of postoperative depression (POD) in recent years has placed a heavy burden on patients' physical and mental health. At this point in time, however, POD pathogenesis remains poorly understood and novel therapeutic strategies are being sought. The present study aimed to clarify esketamine's protective effects and possible mechanisms of action in POD. To this avail, we used an animal model of postoperative depression to analyze behavioral, parameters, plus the inflammatory response in serum and in the medial prefrontal cortex (mPFC). Using immunofluorescence staining, we detected the number of microglia and parvalbumin (PV) in mPFC, and determined changes in neuronal dendritic spine density via Golgi staining. Expression of Iba1, PSD95 and NF-κB was examined by Western blot analysis. Our results show that esketamine can significantly improve depression-like symptoms caused by anesthesia and surgery. In addition, esketamine administration reversed the decrease in the density of PV neurons and restored synaptogenesis in mPFC which had been perturbed by inflammation. The evidence obtained suggests esketamine's anti-inflammatory effects may be mediated by the BDNF/TrkB signaling pathway and possibly by attenuation of the nuclear factor κB (NF-κB) pathway. These data warrant further investigations into the interplay of esketamine, and microglia in the modulation of POD symptomatology.

Differentiation Between Primary Central Nervous System Lymphoma and Atypical Glioblastoma Based on MRI Morphological Feature and Signal Intensity Ratio: A Retrospective Multicenter Study.

OBJECTIVES: To investigate the value of morphological feature and signal intensity ratio (SIR) derived from conventional magnetic resonance imaging (MRI) in distinguishing primary central nervous system lymphoma (PCNSL) from atypical glioblastoma (aGBM). METHODS: Pathology-confirmed PCNSLs (n = 93) or aGBMs (n = 48) from three institutions were retrospectively enrolled and divided into training cohort (n = 98) and test cohort (n = 43). Morphological features and SIRs were compared between PCNSL and aGBM. Using linear discriminant analysis, multiple models were constructed with SIRs and morphological features alone or jointly, and the diagnostic performances were evaluated via receiver operating characteristic (ROC) analysis. Areas under the curves (AUCs) and accuracies (ACCs) of the models were compared with the radiologists' assessment. RESULTS: Incision sign, T2 pseudonecrosis sign, reef sign and peritumoral leukomalacia sign were associated with PCNSL (training and overall cohorts, P < 0.05). Increased T1 ratio, decreased T2 ratio and T2/T1 ratio were predictive of PCNSL (all P < 0.05). ROC analysis showed that combination of morphological features and SIRs achieved the best diagnostic performance for differentiation of PCNSL and aGBM with AUC/ACC of 0.899/0.929 for the training cohort, AUC/ACC of 0.794/0.837 for the test cohort and AUC/ACC of 0.869/0.901 for the overall cohort, respectively. Based on the overall cohort, two radiologists could distinguish PCNSL from aGBM with AUC/ACC of 0.732/0.724 for radiologist A and AUC/ACC of 0.811/0.829 for radiologist B. CONCLUSION: MRI morphological features can help differentiate PCNSL from aGBM. When combined with SIRs, the diagnostic performance was better than that of radiologists' assessment.

[Inhibition effect and mechanism of artemisnin on surgically induced endometriosis].

OBJECTIVE: To evaluate the inhibitory effect of artemisnin on surgically induced endometriosis in rat model and the possible mechanism related to cellular apoptosis and microvascular angiogenesis. METHODS: Surgically induced endometriosis model was established with female rats, and then the rats were divided into four groups: high dose artemisinin [300 mg/(kg x d)), low dose artemisinin [150 mg/(kg x d)], danazol [160 mg/(kg x d)] and solvent control group After daily administration of the above agents for 4 weeks, the rats were sacrificed, then the implant size of ectopic endometrium was measured and appotosis index (AI), Bcl-2 and MVD in ectopic endometrium were evaluated with S-P immunohistochemistry. RESULTS: Compared with solvent control, both artemisinin (high and low quality) and danazol decrease the size of implants significantly (P < 0.05), and there was no significant difference among the three treatment groups. AI of the three treatment groups increased significantly, while Bcl-2 and MVD decreased significantly (P < 0.05). AI of both artemisinin groups were significantly higher than that of danazol group, but Bcl-2 level was lower. CONCLUSION: Artemisnin inhibits surgically induced endometriosis in rats, and the possible mechanism may be related to stimulatation of cellular apoptosis and inhibition of angiogenesis.

Screening and identification of small molecule compounds perturbing mitosis using time-dependent cellular response profiles.

Cellular processes such as cell cycle progression, mitosis, apoptosis, and cell migration are characterized by well-defined events that are modulated as a function of time. Measuring these events in the context of time and its perturbation by small molecule compounds and RNAi can provide mechanistic information about cellular pathways being affected. We have used impedance-based time-dependent cell response profiling (TCRP) to measure and characterize cellular responses to antimitotic compounds or siRNAs. Our findings indicate that small molecule perturbation of mitosis leads to unique TCRP. We have further used this unique TCRP signature to screen 119 595 compound library and identified novel antimitotic compounds based on clustering analysis of the TCRPs. Importantly, 113 of the 117 hit compounds in the TCRP antimitotic cluster were confirmed as antimitotic based on independent assays, thus establishing the robust predictive nature of this profiling approach. In addition, potent and novel agents that induce mitotic arrest either by directly interfering with tubulin polymerization or by other mechanisms were identified. The TCRP approach allows for a practical and unbiased phenotypic profiling and screening tool for small molecule and RNAi perturbation of specific cellular pathways and time resolution of the TCRP approach can serve as a complement for other existing multidimensional profiling approaches.

Luteoloside Exerts Analgesic Effect in a Complete Freund's Adjuvant-Induced Inflammatory Model via Inhibiting Interleukin-1ß Expression and Macrophage/Microglia Activation.

BACKGROUND: Flavonoid monomers are proved to have an anti-inflammatory effect and may also be promising for chronic pain treatment. In the present study, the analgesic effect and the relevant mechanisms of luteoloside, one of the flavonoid monomers, were investigated. METHODS: The analgesic effect of luteoloside was first evaluated in complete Freud's adjuvant induced inflammatory model by von Frey test and Hargreaves test in both male and female mice. The interleukin-1ß levels in plantar tissue, serum, dorsal root ganglion, and the dorsal horn of the spinal cord were determined by enzyme-linked immunosorbent assay or immunofluorescence. The activation of macrophage/microglia was tested by Iba-1 staining. RESULTS: Our data showed that luteoloside exhibited both acute and chronic analgesic phenotypes. Every single dose of luteoloside solution reached the peak transient analgesic effect 2 h after administration and lasted less than 6 h. About 14 consecutive days administration (one dose per day) later, luteoloside showed a sustained analgesic effect which lasted more than 24 h. Celecoxib 20 mg/kg combined with luteoloside 40 mg/kg achieved a similar analgesic effect as celecoxib 40 mg/kg alone. Luteoloside inhibited interleukin-1ß expression in plantar tissue, dorsal root ganglion, the dorsal horn of spinal cord, and serum, after 14 days of continuous administration. Furthermore, our results also showed that the activation of macrophage/microglia in dorsal root ganglions were significantly inhibited 2 h after each single dose in daily luteoloside administration and recovered to a higher level 6 h later. These findings might be involved in the mechanisms of the acute analgesic effect of luteoloside. CONCLUSION: Luteoloside presents an analgesic effect via anti-inflammatory and other mechanisms such as inhibiting the activation of macrophage/microglia.

Crosstalk between cardiomyocytes and noncardiomyocytes is essential to prevent cardiomyocyte apoptosis induced by proteasome inhibition.

Heart is a multi-cellular organ made up of various cell types interacting with each other. Cardiomyocytes may benefit or suffer from crosstalk with noncardiomyocytes in response to diverse kinds of cardiac stresses. Proteasome dysfunction is a common cardiac stress which causes cardiac proteotoxicity and contributes to cardiac diseases such as heart failure and myocardial infarction. The role of crosstalk between cardiomyocytes and noncardiomyocytes in defense of cardiac proteotoxicity remains unknown. Here, we report a cardiomyocyte-specific survival upon proteasome inhibition in a heterogeneous culture consisting of cardiomyocytes and other three major cardiac cell types. Conversely, cardiomyocyte apoptosis is remarkably induced by proteasome inhibition in a homogeneous culture consisting of a majority of cardiomyocytes, demonstrating an indispensable role of noncardiomyocytes in the prevention of cardiomyocyte apoptosis resulting from proteasome inhibition. We further show that cardiomyocytes express brain natriuretic peptide (BNP) as an extracellular molecule in response to proteasome inhibition. Blockade of BNP receptor on noncardiomyocytes significantly exacerbated the cardiomyocyte apoptosis, indicating a paracrine function of cardiomyocyte-released extracellular BNP in activation of a protective feedback from noncardiomyocytes. Finally, we demonstrate that proteasome inhibition-activated transcriptional up-regulation of BNP in cardiomyocytes was associated with the dissociation of repressor element 1 silencing transcription factor (REST)/ histone deacetylase 1 (HDAC1) repressor complex from BNP gene promoter. Consistently, the induction of BNP could be further augmented by the treatment of HDAC inhibitors. We conclude that the crosstalk between cardiomyocytes and noncardiomyocytes plays a crucial role in the protection of cardiomyocytes from proteotoxicity stress, and identify cardiomyocyte-released BNP as a novel paracrine signaling molecule mediating this crosstalk. These findings provide new insights into the key regulators and cardioprotective mechanism in proteasome dysfunction-related cardiac diseases.

[Screening and identification of differentially expressed genes in Beijing fatty and broiler breast muscles].

mRNA differential display reverse-transcripton PCR(DDRT-PCR) was applied to identify differentially expressed genes in Arbor Acres broiler(AA) and Beijing fatty chicken breast muscles in order to find the mechanism which induces the differential gene expression at the molecular level. A total of 7 ESTs were found using reverse Northern dot blot, and all of them were compared with the nucleotide sequences in GenBank database using BLAST. S1 was highly similar to HMGN3; S3 was highly similar to ChEST294a8 with unknown functions; S4 and S5 were highly similar to PGM; S6 and S2 had highly similar nucleotide sequences with unknown functions in nucleotide databases; S7 had no significant similarity with existing genes or ESTs and was regarded as a new EST. The new EST was submitted to GenBank(Accession number: EU594549). This lays a foundation for further study on the mechanism of differential gene expression in Beijing fatty and AA breast muscles.