Effects of dietary multienzymes on the growth performance, digestive enzyme activity, nutrient digestibility, excreta noxious gas emission, and nutrient transporter gene expression in white feather broilers.

Adding multienzymes to poultry feed rations is recognized as a nutritional strategy aimed at improving poultry performance and health status. Nonetheless, some literatures present an ongoing debate about the extent of multienzymes beneficial impact on poultry growth performance. This study aimed to explore the impacts of dietary multienzyme supplementation on broilers, focusing specifically on growth performance, carcass characteristics, apparent nutrient digestibility, excreta noxious gas emission, and intestinal nutrient transporter gene expression. A total of 3,200 broilers were randomly assigned to five groups (eight replicates per treatment group) and treated with the following: normal control (CON), CON + 100 g/t multienzyme (ME100), CON + 150 g/t multienzyme (ME150), CON + 200 g/t multienzyme (ME200), and CON + 250 g/t multienzyme (ME250). Supplementing with multienzymes significantly influenced the feed conversion rate (linear, P = 0.007; quadratic, P = 0.024) and the European broiler index (linear, P = 0.004; quadratic, P = 0.016) in broilers. Dietary multienzymes significantly influenced apparent metabolizable energy (quadratic, P = 0.015) and neutral detergent fiber (quadratic, P < 0.001). Moreover, multienzyme supplementation in the diet also decreased the emission of ammonia (linear, P = 0.001; quadratic, P = 0.006) and hydrogen sulfide (quadratic, P = 0.006) in the excreta. In addition, dietary multi-enzyme notably elevated (P < 0.05) the mRNA expression of nutrient transporter genes, including peptide transporter 1 (PePT1), Na-dependent neutral amino acid transporter (B0AT), glucose transporter 2 (GLUT2), and fatty acid binding protein1 (FABP1). These findings suggest that dietary supplementation with multienzymes can improve the efficiency of feed utilization, and the digestion and absorption of nutrients and reduce excreta gas emission. Furthermore, this study provides a theoretical basis for advancing the use of multienzymes in broiler production.

Multienzyme additives are increasingly used in animal feed, primarily to enhance growth performance and nutrient digestibility. This study focused on the effects of multienzyme additives (xylanase, mannanase, cellulase, arabinofuranosidase, ferulic acid esterase, amylase, and protease) on various aspects of broilers, including growth performance, carcass characteristics, digestive enzyme activities, apparent nutrient digestibility, excreta noxious gas emission, and intestinal nutrient transporter gene expression. The inclusion of multienzymes in the diet was found to significantly increase the weight of breast muscle in broilers. Additionally, it led to a notable decrease in the viscosity of the fecal and jejunal digesta. Furthermore, the present study revealed an increase in the mRNA expression of key nutrient transporters­peptide transporter 1 (PePT1), Na-dependent neutral amino acid transporter (B0AT), and fatty acid binding protein 1 (FABP1), in the intestine of broilers. These findings indicate that dietary multienzymes enhance the efficiency of feed nutrient digestion and absorption in broilers.

Dietary novel alkaline protease from Bacillus licheniformis improves broiler meat nutritional value and modulates intestinal microbiota and metabolites.

BACKGROUND: Different types of exogenous protease supplements have a positive impact on animal performance, but their effects on the nutritional value of meat and the gut microbial community of broilers have not been extensively studied. The objective of this investigation was to determine the impact of supplementation with a novel alkaline protease derived from Bacillus licheniformis (at doses of 0, 100, 200, 300, and 400 g/t) on the fatty acid and amino acid profiles, inosine monophosphate (IMP) levels, total volatile basic nitrogen (TVB-N) content found within the breast muscle, as well as the impact on the cecal microbiota and metabolites. RESULTS: Supplementation with 200-400 g/t of the novel protease resulted in a significant elevation in the concentration of essential amino acids (P < 0.001), flavor amino acids (P < 0.001), and total protein (P = 0.013) within the breast muscle. Results derived from the 16S rRNA sequencing and untargeted metabolomics analysis of the cecal content revealed that the novel protease reshaped the cecal microbial and metabolite profiles. In particular, it led to increased relative abundances of Bacteroides, Lactobacillus, Alistipes, and Eubacterium, while simultaneously causing a reduction in the metabolites of D-lactic acid and malonic acid. Moreover, correlation analyses unveiled significant relationships between distinct microbes and metabolites with the contents of IMP, fatty acids, and amino acids in the broiler's breast muscle. CONCLUSION: In summary, the novel protease regulated the intestinal microbial community and metabolism, thereby inducing changes in the compositions of fatty acids and amino acids profiles, as well as IMP levels in broiler meat. These alterations significantly contributed to the enhancement of the nutritional value and flavor of the meat.

Effect of a novel alkaline protease from Bacillus licheniformis on growth performance, carcass characteristics, meat quality, antioxidant capacity, and intestinal morphology of white feather broilers.

BACKGROUND: The present study was conducted to investigate the effects of dietary novel alkaline protease from Bacillus licheniformis on the growth performance, meat quality, antioxidant status and intestinal morphology of broilers. In total, 4000 broilers were randomly assigned into five groups and treated with normal control, normal control + 100 mg kg-1 protease, normal control + 200 mg kg-1 protease, normal control + 300 mg kg-1 protease and normal control + 400 mg kg-1 protease. RESULTS: Supplementing protease impacted final body weight (linear, P = 0.003; quadratic, P = 0.006) and decreased feed conversion rate (linear, P = 0.036) in broilers. Moreover, dietary protease significantly increased breast muscle rate (linear, P = 0.005; quadratic, P = 0.021) and decreased drip loss (linear, P < 0.001; quadratic, P < 0.001). In addition, dietary protease notably increased protein digestibility (linear, P = 0.001; quadratic, P = 0.006) and trypsin activity (linear, P = 0.002; quadratic, P = 0.009) in jejunum. Light microscopy revealed that the jejunum villi in the 300 mg kg-1 and 400 mg kg-1 groups exhibited greater height and a denser arrangement compared to those in the control group. The addition of protease decreased malondialdehyde content (linear, P < 0.001; quadratic, P < 0.001) and increased total antioxidant capacity (linear, P = 0.001; quadratic, P < 0.001) in pectoral muscles. CONCLUSION: The results of the present study suggest that dietary novel alkaline protease from B. licheniformis improved growth performance by affecting trypsin activity, protein digestibility, antioxidant capacity and intestinal health. © 2024 Society of Chemical Industry.

[Teaching reform and practice of 'Production Internship' course for biotechnology specialty from the perspective of training application-oriented talents].

Production internship is an important teaching tache for undergraduate students to carry out engineering training by using professional skills, and it is a key starting point for fostering application-oriented talents in biotechnology. The Course Group of 'production internship of biotechnology majors' of Binzhou University is investigating application-oriented transformation for local regular colleges and universities, as well as fostering high-level application-oriented talents. By taking green fluorescent protein (GFP) polyclonal antibody as an example, the reform and practice on teaching content, teaching mode, assessment method, continuous improvement of curriculum were carried out. Moreover, the characteristics of the Yellow River Delta-Binzhou Biotechnology & Pharmaceutical Industrial Cluster were taken into account to intensify academic-enterprise cooperation. On one hand, this Course Group designed and rearranged the course contents, carried out essential training through online resources and platforms such as virtual simulation, and recorded, tracked and monitored the progress of production internship through practical testing and software platforms like 'Alumni State'. On the other hand, this Course Group established a practice-and application-oriented assessment method in the process of production internship and a dual evaluation model for continuous improvement. These reform and practices have promoted the training of application-oriented talents in biotechnology, and may serve as a reference for similar courses.

Recombinant human ß-defensin130 inhibited the growth of foodborne bacteria through membrane disruption and exerted anti-inflammatory activity.

Foodborne pathogens causing food poisoning and infections are detrimental to human health, and the abuse of antibiotics induced severe antibiotic resistance in past decades. Thus, it is urgent to develop new antimicrobial agents. In the current study, human ß-defensin 130 (hBD130), which is an antimicrobial peptide identified in human macrophages in 2017, was initially produced in Pichia pastoris. The purified hBD130 demonstrated broad bactericidal spectrum against foodborne pathogens through a membrane disruption, with concentrations ranging from 10 to 45 µg/mL. Moreover, hBD130 showed a low hemolytic effect and nearly no cytotoxicity to mammalian cells with a dosage of 400 µg/mL. In addition, the secretion amounts and mRNA levels of NO, IL-6, IL-1ß, and TNF-α in LPS-induced mouse macrophage were significantly decreased with 1 mg/mL of hBD130. Taken together, these results showed that hBD130 is a promising antimicrobial agent to treat foodborne bacterial infections and inflammation. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-022-01087-y.

Antibacterial activity of antimicrobial peptide gcDefb1 against foodborne pathogenic bacteria and its application in pork storage.

Pork and its products are preferably contaminated by bacteria; thus, it is essential to develop low-cost, high-efficiency and biologically safe preservatives to prevent the growth of bacteria during storage. In the current study, grass carp ß-defensin 1 (gcDefb1) was produced and purified from Pichia pastoris through the heterologous expression method. The in vitro antimicrobial assay demonstrated that yeast-derived gcDefb1 possesses a broad antibacterial spectrum, including both Gram-positive and -negative bacteria, and the MIC values against Escherichia coli ATCC 25,922 were as low as 30 µg/mL and showed no cytotoxicity or hemolytic activity. The bactericidal rate of gcDefb1 was less than 60 min by disrupting the cell membranes, and it inhibited the formation of bacterial biofilms. Moreover, gcDefb1 was used as a biopreservative for pork storage, indicating that the physicochemical and sensory qualities were improved. This study provides an efficient method to prepare and utilize gcDefb1 as a novel biopreservative. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-022-01060-9.

A method to predict electric vehicles' market penetration as well as its impact on energy saving and CO2 mitigation.

INTRODUCTION: Although forecasting electric vehicles' growth in China was frequently reported in the literature, predicting electric vehicles market penetration as well as corresponding energy saving and carbon dioxide mitigation potential in a more suitable method is not well understood. METHODS: This study chose the double species model to predict electric vehicles' growth trajectory under mutually competitive conditions between electric vehicles and internal combustion engine vehicles. For comparison, it set two scenarios: with 200 and 300 vehicles per thousand persons at 2050. To give details on energy saving and carbon dioxide mitigation potential induced by electric vehicles' market penetration, it further divided electric vehicles into five subgroups and internal combustion engine vehicles into seven subgroups, therein forming respective measurement formulas. RESULTS: This paper solved the double species model and thus got its analytical formula. Then it employed the analytical formula to conduct an empirical study on electric vehicles market penetration in China from year 2010 to 2050. Under scenario 300, electric vehicles growth trajectory will emerge a quick growth stage during 2021-2035, thereafter keeping near invariant till 2050. Meanwhile, current internal combustion engine vehicles' quick growth will continue up to 2027, then holding constant during 2028-2040, afterwards following a 10-year slowdown period. Scenario 200 has similar features, but a 2-year delay for electric vehicles and a 5-year lead time for internal combustion engine vehicles were found. On average, scenario 300 will save 114.4 Mt oil and 111.5 Mt carbon dioxide emissions, and scenario 200 will save 77.1 Mt oil and 73.4 Mt carbon dioxide emissions each year. Beyond 2032, annual 50.0% of road transport consumed oil and 18.6% of carbon dioxide emissions from this sector will be saved under scenario 300. DISCUSSION: Compared with scenario 200, scenario 300 was more suitable to predict electric vehicle market penetration in China. In the short-term electric vehicle penetration only brings about trivial effects, while in the long-term it will contribute a lot to both energy security and carbon dioxide mitigation. The contribution of this article provided a more suitable methodology for predicting electric vehicle market penetration, simulated two coupled trajectories of electric vehicles and internal combustion engine vehicles, and discussed relative energy-saving and climate effects from 2010 to 2050.

Incidence and PD-L1 Expression of MET 14 Skipping in Chinese Population: A Non-Selective NSCLC Cohort Study Using RNA-Based Sequencing.

BACKGROUND: Mesenchymal-epithelial transition (MET) exon14 skipping mutations represent a clinically unique molecular subtype of NSCLC. The prevalence rates of MET exon 14 skipping in lung adenocarcinoma (ADC) range from 0.9% to 4.0% in Asian populations. Since some somatic variants that do not encompass the MET exon 14 splice sites might also induce MET exon 14 skipping, the RNA-based sequencing is speculated as the most accurate method for detecting exon 14 skipping. PATIENTS AND METHODS: A total of 951 NSCLC patients from two hospitals were enrolled in this study. MET exon14 skipping was detected using RNA-based next-generation sequencing (NGS). Also, immunohistochemistry (IHC) was performed in 405 samples simultaneously. RESULTS: The overall estimated prevalence of MET exon 14 skipping was approximately 1.8% in ADCs and 1.7% in NSCLCs. The detection rate of MET exon 14 skipping from surgical resection specimen was 2.3% in NSCLCs and 2.0% in ADCs. The MET exon 14 skipping was identified in 6.6% of EGFR/KRAS/ALK/ROS1/RET-negative ADCs. Additionally, PD-L1 was found to be highly expressed in NSCLC patients harboring MET exon 14 skipping (P<0.01). CONCLUSION: The prevalence of MET exon14 skipping in lung ADCs in the East Asian population was similar to that of the Western population as assessed by RNA-based NGS. The NSCLC patients with MET exon 14 skipping were older than those with other oncogenic driver mutations, such as EGFR, ALK, and ROS1. In addition, PD-L1 was highly expressed in NSCLC patients with MET exon 14 skipping.

Simultaneous Determination of Eight Active Compounds in Baitouweng Decotion and its Single Herbs.

Baitouweng Decotion (BD) is a famous traditional Chinese medicinal prescription, which is composed of Pulsatilla chinensis (bunge) regel, Coptis chinensis franch., Phellodendron chinense and Cortex Fraxini. In this study, a simple and sensitive high performance liquid chromatography coupled with ultraviolet detection method was established for the simultaneous determination of eight marker compounds including Esculin, Fraxin, Esculetin, Fraxetin, Columbamine, Coptisine, Palmatine Chloride and Berberine hydrochloride in BD, the single herbs and their negative controls. The chromatographic separation was performed using an Agilent Eclipse XDB-C18 column with a gradient elution system of acetonitrile and 0.1% phosphoric acid (contained 0.2% triethylamine) solution at a flow rate of 0.8 mL/min. The results demonstrated that the validated method was simple, reliable and successfully applied to evaluate the selected compounds in water extraction (BDW) and ethanol extraction (BDE) of BD, the single herbs and their negative control for quality control. Moreover, the experimental data showed that the contents of the major active components detected in BDE were significantly higher than those in the BDW, while the BDW had several peaks BDE without. The paper also suggested a method to extract Fraxin, Esculin, Fraxetin, Esculetin and Berberine from Baitouweng Decotion more effectively.

Prevalence of ROS1 fusion in Chinese patients with non-small cell lung cancer.

BACKGROUND: The study was conducted to investigate the clinicopathological features and prevalence of ROS1 gene fusion in Chinese patients with non-small cell lung cancer (NSCLC). METHODS: The presence of ROS1 fusion was assessed by quantitative real-time PCR. Associations between ROS1 fusion and clinical characteristics were analyzed. RESULTS: In total, 6066 patients with pathologically confirmed NSCLC and ROS1 fusion test results were enrolled. The average age was 60.89 ± 10.60 years and fusion was detected in 157 (2.59%) patients. Fusion frequency was significantly correlated with age, gender, smoking status (all P < 0.001), pathology type (P = 0.017), and lymph node metastasis stage (P = 0.027). ROS1 fusion-positive patients were significantly younger (55.68 ± 11.34 vs. negative 61.02 ± 10.44 years; P < 0.01). Fusion frequency was higher in women (3.71% vs. men 1.81%), never-smokers (3.33% vs. smokers 1.21%), and patients with adenocarcinoma (2.77% vs. squamous lung cancer 0.93%) and at advanced node stages (1.31%, 1.40%, 2.07%, and 3.23% for N0, N1, N2, and N3, respectively). No significant correlation between ROS1 fusion status and pathological stage was found in subgroups classified by pathological, tumor, or metastasis stage (P > 0.05). Age, smoking status, and lymph node stage were statistically significantly correlated with ROS1 fusion frequency (all P < 0.05); gender and pathology type were not significantly correlated with ROS1 fusion status after adjusting for smoking status. CONCLUSION: An overall ROS1 fusion frequency of 2.59% was confirmed in this study. ROS1 fusion was more prevalent among younger patients, never-smokers, and those at advanced node stages.

Cost-effectiveness of ALK testing and first-line crizotinib therapy for non-small-cell lung cancer in China.

INTRODUCTION: Anaplastic lymphoma kinase (ALK) rearrangement gene testing is used increasingly to identify patients with advanced non-small-cell lung cancer (NSCLC) who are most likely to benefit from crizotinib. This study was to evaluate the cost-effectiveness of the ALK tests followed by crizotinib compared to the standard chemotherapy in advanced NSCLC from the Chinese healthcare system perspective. METHODS: A 10-year Markov model was constructed to compare the costs and quality-adjusted life-years (QALYs) of crizotinib with standard chemotherapy, guided by the ALK rearrangement tests: next-generation sequencing (NGS) panel tests and multiplex polymerase chain reaction (PCR) testing. The health states included progression-free survival (PFS), progressed survival, and death. The costs examined included cost of drugs (pemetrexed, standard chemotherapy, salvage chemotherapy, and crizotinib), follow-up, palliative care, supportive care, severe adverse events, and ALK rearrangement testing. RESULTS: Under Patient Assistance Program (PAP), the model demonstrated that the patients using NGS panel tests spent US $31,388 and gained 0.780 QALYs, whereas patients using multiplex PCR spent US $31,362 and gained 0.780 QALYs, respectively. The incremental cost-effectiveness ratios of crizotinib with PAP compared to the control strategy were projected at $14,384 (NGS) and $13,740 (multiplex PCR) per QALY gained, respectively. Sensitivity analyses showed the utility of PFS and the costs of crizotinib and pemetrexed were the most impactful factors on the model outcomes. The results were robust to changes in all parameters. CONCLUSION: ALK-rearrangement test positive followed by crizotinib may be cost-effective compared to standard chemotherapy from the Chinese healthcare system perspective when PAP was available.

Comprehensive Characterization of Oncogenic Drivers in Asian Lung Adenocarcinoma.

INTRODUCTION: The incidence rate of lung adenocarcinoma (LUAD), the predominant histological subtype of lung cancer, is elevated in Asians, particularly in female nonsmokers. The mutation patterns in LUAD in Asians might be distinct from those in LUAD in whites. METHODS: We profiled 271 resected LUAD tumors (mainly stage I) to characterize the genomic landscape of LUAD in Asians with a focus on female nonsmokers. RESULTS: Mutations in EGFR, KRAS, erb-b2 receptor tyrosine kinase 2 gene (ERBB2), and BRAF; gene fusions involving anaplastic lymphoma receptor tyrosine kinase gene (ALK), ROS1, and ret proto-oncogene (RET); and Met Proto-Oncogene Tyrosine Kinase (MET) exon 14 skipping were the major drivers in LUAD in Asians, exhibiting mutually exclusive and differing prevalence from those reported in studies of LUAD in non-Asians. In addition, we identified a novel mutational signature of XNX (the mutated base N in the middle flanked by two identical bases at the 5' and 3' positions) that was overrepresented in LUAD tumors in nonsmokers and negatively correlated with the overall mutational frequency. CONCLUSIONS: In this cohort, approximately 85% of individuals have known driver mutations (EGFR 59.4%, KRAS 7.4%, ALK 7.4%, ERBB2 2.6%, ROS1 2.2%, RET 2.2%, MET 1.8%, BRAF 1.1%, and NRAS 0.4%). Seventy percent of smokers and 90% of nonsmokers had defined oncogenic drivers matching the U.S. Food and Drug Administration-approved targeted therapies.

Structure and antigenicity analysis of the IgG gene for Nyctereutes procyonoides.

OBJECTIVE: Nyctereutes procyonoides immunoglobulin G (IgG) gene is partially cloned. MATERIAL AND METHODS: In order to obtain a certain length (966bp) of Nyctereutes procyonoides immunoglobulin G (IgG), two pairs of primers are designed according to the conserved nucleotide sequence of canine (GenBank:AF354265, AF354265, AF354266, AF354267) and mink (GenBank: L07789). Using Bioinformatics technology and Western-blot to analyze antigenicity of Nyctereutes procyonoides IgG-B gene. RESULTS: The homology for nucleotide sequence of IgG between Nyctereutes procyonoides and canine (IgG A, IgG B, IgG C, IgG D), mink, Homo sapiens, Oryctolagus cuniculus, Mus musculus, Anas platyrhynchos and gallus were respectively (88.1%, 93.6%, 85.4%, 87.2%), 83.7%, 74.8%, 71.8%, 69.2%, 51.6%, 48.4%. It can be seen that there was high homology of aminoacid sequence between IgG of Nyctereutes procyonoides and IgG (A, B, C, D) of canine. And the serum antibody of Nyctereutes procyonoides had obviously cross-reaction with HRP conjugated rabbit anti-dog IgG, compared with those of canine, oryctolagus cuniculus, mus musculus, mink, gallus. CONCLUSIONS: We successfully got Nyctereutes procyonoides immuneglobulin G (IgG) gene (Gen- Bank: KM010191). There is the closest ties of consanguinity of IgG exist between Nyctereutes procyonoides and canine among the mammal through the genetic evolution. The detection and treament of canine distemper can be used on Nyctereutes procyonoides.

Immune function of Chinese formula Qingwen Baidu granule in broilers.

This study was to investigate the effects of Qingwen Baidu granules on the antibody level, immune organ index and the lymphocyte transformation of broilers. Hy-line variety white cocks of 30 days were used to evaluate the antibody titer of Newcastle Disease in each serum group, and MTT method was used to determine the T lymphocyte proliferation, and organ weighing methods to measure the immune organ index 21 days after immunization. The results showed that Qingwen Baidu granules could prolong the residue time in the body, improve the lymphocyte conversion ratio, increase the bursa, thymus and spleen index and promote immune organ development. These results suggested that Qingwen Baidu granules could improve the serum Newcastle disease antibody level, improve peripheral blood lymphocyte proliferation, enhance the cellular immune function, and elevate the immune organ index and growth, in order to raise the immune function in chicken. The above demonstrates that the Qingwen Baidu granules have significant effects on the cytoimmunity and humoral immunity, and the potentiation of the immune function in broilers.

Histone methyltransferase SETDB1 regulates liver cancer cell growth through methylation of p53.

SETDB1 is a histone H3K9 methyltransferase that has a critical role in early development. It is located within a melanoma susceptibility locus and facilitates melanoma formation. However, the mechanism by which SETDB1 regulates tumorigenesis remains unknown. Here we report the molecular interplay between SETDB1 and the well-known hotspot gain-of-function (GOF) TP53 R249S mutation. We show that in hepatocellular carcinoma (HCC) SETDB1 is overexpressed with moderate copy number gain, and GOF TP53 mutations including R249S associate with this overexpression. Inactivation of SETDB1 in HCC cell lines bearing the R249S mutation suppresses cell growth. The TP53 mutation status renders cancer cells dependent on SETDB1. Moreover, SETDB1 forms a complex with p53 and catalyses p53K370 di-methylation. SETDB1 attenuation reduces the p53K370me2 level, which subsequently leads to increased recognition and degradation of p53 by MDM2. Together, we provide both genetic and biochemical evidence for a mechanism by which SETDB1 regulates cancer cell growth via methylation of p53.

Altered baseline brain activity in experts measured by amplitude of low frequency fluctuations (ALFF): a resting state fMRI study using expertise model of acupuncturists.

It is well established that expertise modulates evoked brain activity in response to specific stimuli. Recently, researchers have begun to investigate how expertise influences the resting brain. Among these studies, most focused on the connectivity features within/across regions, i.e., connectivity patterns/strength. However, little concern has been given to a more fundamental issue whether or not expertise modulates baseline brain activity. We investigated this question using amplitude of low-frequency (<0.08 Hz) fluctuation (ALFF) as the metric of brain activity and a novel expertise model, i.e., acupuncturists, due to their robust proficiency in tactile perception and emotion regulation. After the psychophysical and behavioral expertise screening procedure, 23 acupuncturists and 23 matched non-acupuncturists (NA) were enrolled. Our results explicated higher ALFF for acupuncturists in the left ventral medial prefrontal cortex (VMPFC) and the contralateral hand representation of the primary somatosensory area (SI) (corrected for multiple comparisons). Additionally, ALFF of VMPFC was negatively correlated with the outcomes of the emotion regulation task (corrected for multiple comparisons). We suggest that our study may reveal a novel connection between the neuroplasticity mechanism and resting state activity, which would upgrade our understanding of the central mechanism of learning. Furthermore, by showing that expertise can affect the baseline brain activity as indicated by ALFF, our findings may have profound implication for functional neuroimaging studies especially those involving expert models, in that difference in baseline brain activity may either smear the spatial pattern of activations for task data or introduce biased results into connectivity-based analysis for resting data.

Development of a miR-26 companion diagnostic test for adjuvant interferon-alpha therapy in hepatocellular carcinoma.

BACKGROUND & AIMS: Adjuvant therapies for hepatocellular carcinoma (HCC) such as interferon-alpha are effective only in a subset of patients. Previously we found that HCC patients with low level of miR-26 have survival benefits from interferon-alpha. The purpose of this study is to develop a standardized miR-26 diagnostic test (referred as MIR26-DX) to assist identification of candidate HCC patients for adjuvant interferon-alpha therapy. METHODS: We developed a multiplex reverse-transcription quantitative polymerase-chain-reaction assay to determine the levels of two HCC-related miR-26 transcripts along with six small RNA reference transcripts. We evaluated archived paraffin-embedded tissues from three cohorts of HCC patients (n=248) who underwent radical resection at three different clinical centers. Fifty-two percent of them underwent adjuvant interferon-alpha therapy. We used Cox-Mantel log-rank test to evaluate patient survival. RESULTS: We found that the multiplexing assay was stable and reproducible regardless of differences in sample preparations and operators. We developed a matrix template and a scoring algorithm based on a training cohort (n=129) to assign HCC patients, and then applied the template in two test cohorts (n=119). The proportions of HCC patients assigned as low miR-26 by this algorithm were 68, 4, and 63 percent in the training cohort and two test cohorts, respectively. Consistently, HCC with low miR-26 had a favorable response to interferon-alpha with improved median overall survival (≥3 year). CONCLUSIONS: MIR26-DX is a simple and reliable companion diagnostic test to select HCC patients for adjuvant interferon-alpha therapy.

Multi-microarray identifies lower AQP9 expression in adjuvant chemotherapy nonresponders with stage III colorectal cancer.

Approximately 25% of stage III colorectal cancer patients do not benefit from standard adjuvant chemotherapies. To identify biomarkers for nonresponders, fresh-frozen tissues of 19 nonresponders and 16 responders were analyzed with gene expression and microRNA arrays. Fifty-nine genes and 17 miRNAs were differentially expressed by at least two folds. AQP9, SATB2, and WIF1 were simultaneously lower expressed in the nonresponders and modulated by the differentially expressed miRNAs. RT-PCR validated the differential expression of AQP9 (p=0.035). In conclusion, lower AQP9 gene expression is related with non-response to adjuvant chemotherapy, showing potential as predictive marker and therapeutic target.

Nurr1 regulates Top IIß and functions in axon genesis of mesencephalic dopaminergic neurons.

BACKGROUND: NURR1 (also named as NR4A2) is a member of the steroid/thyroid hormone receptor family, which can bind to DNA and modulate expression of target genes. Previous studies have shown that NURR1 is essential for the nigral dopaminergic neuron phenotype and function maintenance, and the defects of the gene are possibly associated with Parkinson's disease (PD). RESULTS: In this study, we used new born Nurr1 knock-out mice combined with Affymetrix genechip technology and real time polymerase chain reaction (PCR) to identify Nurr1 regulated genes, which led to the discovery of several transcripts differentially expressed in the nigro-striatal pathway of Nurr1 knock-out mice. We found that an axon genesis gene called Topoisomerase IIß (Top IIß) was down-regulated in Nurr1 knock-out mice and we identified two functional NURR1 binding sites in the proximal Top IIß promoter. While in Top IIß null mice, we saw a significant loss of dopaminergic neurons in the substantial nigra and lack of neurites along the nigro-striatal pathway. Using specific TOP II antagonist ICRF-193 or Top IIß siRNA in the primary cultures of ventral mesencephalic (VM) neurons, we documented that suppression of TOP IIß expression resulted in VM neurites shortening and growth cones collapsing. Furthermore, microinjection of ICRF-193 into the mouse medial forebrain bundle (MFB) led to the loss of nigro-striatal projection. CONCLUSION: Taken together, our findings suggest that Top IIß might be a down-stream target of Nurr1, which might influence the processes of axon genesis in dopaminergic neurons via the regulation of TOP IIß expression. The Nurr1-Top IIß interaction may shed light on the pathologic role of Nurr1 defect in the nigro-striatal pathway deficiency associated with PD.

Involvement of histone acetylation of Sox17 and Foxa2 promoters during mouse definitive endoderm differentiation revealed by microRNA profiling.

Generation of hepatocyte from embryonic stem cells (ESCs) holds great promise for hepatocyte replacement therapy to treat liver diseases. Achieving high efficiency of directed differentiation of ESCs to hepatocyte is of critical importance. Previously, Wnt3a has been reported to promote Activin A-induced human definitive endoderm (DE) differentiation, the early stage of hepatocyte differentiation. However, the underlying molecular mechanisms are not clear. Growing evidence demonstrated that microRNAs (miRNAs) are key regulators involved in various important biological processes including the regulation of stem cell differentiation. In the present study, we profiled genome wide miRNA expression during Wnt3a and Activin A induced mouse DE differentiation. We uncovered distinct miRNA expression patterns during DE differentiation with the identification of a subset of miRNAs whose expression is synergistically regulated by Wnt3a/Activin A treatment at different stages of DE differentiation. Forced expression of a pool of such synergistically regulated miRNAs alone could partially promote DE differentiation, indicating a regulatory role of them. Using TargetScan and GeneGO pathway analyses, the synergistically regulated miRNAs are predicted to regulate key pathways involved in DE differentiation; among them includes the regulation of histone acetylation. Consistently, Wnt3a and Activin A treatment increased global histone acetylation which can be partially mimicked by over expression of the pooled miRNAs. Chromatin IP (ChIP) experiments demonstrated that the promoter regions of Sox17 and Foxa2 are subjected to histone acetylation regulation. Administration of Hdac inhibitors greatly augmented DE differentiation. Our data uncovered a novel epigenetic mechanism of Wnt3a and Activin A induced DE differentiation, whereby the treatment of growth factors induced histone acetylation at least in part by the regulation of miRNA expression.