[Silence of circBANP increases radiosensitivity of colorectal cancer cells and inhibits growth of subcutaneous xenografts by up-regulating miR-338-3p expression].

Objective: To investigate the effect of circBANP on radiosensitivity of colorectal cancer cells and subcutaneous transplanted tumor in nude mice and its potential molecular mechanism. Methods: The carcinoma and adjacent normal mucosal tissues of 20 patients with colorectal cancer who were surgically resected in Henan People's Hospital from January 2018 to January 2019 were selected. The radio-resistant colorectal cancer cell LoVo/R was established. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the expressions of circBANP and miR-338-3p. The radiation sensitivity was determined by cell clone formation experiment. Cell vitality was detected by using methyl thiazolyl tetrazolium (MTT). The expressions of autophagy-related protein microtubule-associated protein light chain 3 (LC3) and p62 were detected by western blot. The fluorescence intensity of LC3 in cells was detected by immunofluorescence assay. The downstream microRNAs (miRNAs) of circBANP were predicted by Circular RNA Interactome website and further verified by dual luciferase reporter gene assay. The transplanted tumor model of LoVo/R cells in nude mice was established, and the effect of circBANP on the growth of transplanted tumor after radiation was observed. Results: The expression levels of circBANP and miR-338-3p in colorectal cancer tissues were 3.21+ 0.29 and 0.47+ 0.04, respectively, which were significantly higher than 1.00+ 0.07 and 1.00+ 0.05 in adjacent tissues (P<0.05). The circBANP expression level of LoVo/R cells was 3.21±0.34, higher than 1.00±0.07 of LoVo cells (P<0.05), and the expression level of miR-338-3p of LoVo/R cells was 0.33±0.04, lower than 1.00±0.08 of LoVo cells (P<0.05). After 4 Gy irradiation, compared with the control group, the viability of LoVo/R cells in the circBANP silencing group [(34±4)% vs (62±6)%, P<0.05], the cell survival fraction (0.07±0.02 vs 0.27±0.04, P<0.05) were decreased, and the radiation sensitization ratio was 1.843, the expression of LC3Ⅱ/Ⅰin LoVo/R cells increased while p62 expression decreased, the cell autophagy was observed. Autophagy inhibitor chloroquine reversed the increased expression of LC3Ⅱ/Ⅰ and inhibited expression of p62 in LoVo/R cells induced by radiation, and promoted the suppression of cell viability and survival induced by radiation, the radiotherapy sensitization ratio was 1.780. Compared with control group after 4 Gy irradiation, the relative fluorescence intensity of LC3 in circBANP silencing LoVo/R cells decreased (0.11±0.01 vs 1.00±0.12, P<0.05), the expression of LC3-Ⅱ/Ⅰdecreased (1.25±0.13 vs 3.84±0.39, P<0.05) while p62 expression increased (2.76±0.29 vs 1.00±0.08, P<0.05). As predicted by Circular RNA Interactome website and confirmed by double luciferase reporter gene assay, miR-338-3p was the target gene of circBANP. The relative fluorescence intensity of LC3 in circBANP silencing + anti-miR-338-3p + 4 Gy group increased (7.32±0.72 vs 1.00±0.09, P<0.05), the expression level of LC3-Ⅱ/Ⅰ increased (4.13±0.43 vs 2.31±0.23, P<0.05) while p62 expression decreased (0.34±0.03 and 1.00±0.11, P<0.05), the radiotherapy sensitization ratio was 0.596. Nude mice subcutaneously transplanted tumor experiment showed that the tumor volume and weight of circBANP silencing group on 13, 16, 19, 22, 25, 28, and 31 days were lower than those of control group (P<0.05), while the tumor volume and weight of circBANP silencing + anti-miR-338-3p group on days of 13, 16, 19, 22, 25, 28 and 31 after inoculated were higher than those of circBANP+ anti-miR-NC group (P<0.05). Conclusions: CircBANP can regulate the radiosensitivity of colorectal cancer cells by regulating the expression of miR-338-3p, and affect the growth of transplanted tumor in nude mice. CircBANP may be a potential target for enhancing radiosensitivity of colorectal cancer cells.

[Effects of silencing circRNA ABCB10 expression on biological properties of colorectal cancer cells].

Objective: To investigate the expression of circular ribonucleic acid ABCB10 (circABCB10) in colorectal cancer tissues and cells and its effects on cell biological behavior, radiosensitivity and growth of subcutaneous xenografts. Methods: The tumor tissue and adjacent tissue from colorectal cancer patients treated in Henan People's Hospital were collected from January 2018 to December 2018. Quantitative polymerase chain reaction (qPCR) was used to detect the expressions of circABCB10 and miR-217, cell viability was detected by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H tetrazolium bromide (MTT), cell apoptosis rate was detected by flow cytometry, cell migration and invasion were detected by Transwell method, cell radiosensitivity was detected by colony formation assay. The downstream miRNAs of circABCB10 were predicted by Circular RNA Interactome and verified by the dual luciferase reporter gene experiment. The effect of circABCB10 on the growth of transplanted tumor was examined in nude mice. Results: The expression level of circABCB10 mRNA in colorectal cancer tissues was (3.97±2.12), higher than (1.13±0.64) in adjacent tissues (P<0.05). The expression level of circABCB10 mRNA in FHC cells was (1.00±0.09), lower than that (4.53±0.44) in SW480, (3.12±0.32) in HCT116 and (3.51±0.36) in HT29 cells, respectively (all P<0.05). The MTT results showed that the absorbance values of SW480 cells in si-circABCB10-1 group at 48 and 72 hours after transfection were (0.36±0.04) and (0.43±0.04), lower than (0.48±0.05) and (0.82±0.08) in circ-negative control (NC) group, respectively (all P<0.05). The number of migrating cells and invasive cells in si-circABCB10-1 group were (45±8) and (34±7), lower than (106±21) and (84±15) in circ-NC group, respectively (all P<0.01). The radiosensitization ratio was 1.632. The results of subcutaneous transplantation assay showed that the tumor volume and tumor weight of the si-circABCB10-1 group were significantly lower than circ-NC group after 8 days of inoculation ( all P<0.05). MiR-217 is a target gene of circABCB10. Inhibition of miR-217 reversed the inhibitory effect of circABCB10 silencing on cell proliferation, migration, invasion and subcutaneous xenograft growth in nude mice and the radiosensitization activity. Conclusion: Silence of circABCB10 can up-regulate the expression of miR-217 to inhibit the proliferation, migration, invasion and growth of subcutaneous xenografts and increase the radiosensitivity of SW480 cells, which reveals the underlying molecular mechanism of colorectal cancer progression and provides a new sensitizing target for clinical radiotherapy of colorectal cancer.

[The mechanism of circ_0023990/miR-873-5p/ANXA2 axis regulating radiosensitivity and development of thyroid carcinoma].

Objective: To explore the effect and possible mechanism of circ_0023990 on the radiosensitivity of thyroid cancer cells. Methods: qRT-PCR was used to detect the expression of circ_0023990 in the cancer tissues of 55 patients with thyroid cancer and thyroid cancer cell lines (TPC-1, KTC-1, FTC-133 and CAL-62), and the relationship between the expression of circ_0023990 in cancer tissues and the clinical characteristics of the patients were analyzed. Thyroid cancer cells TPC-1 and KTC-1 were divided into sh-circ_0023990 group, sh-NC group, sh-circ_0023990+anti-miR-873-5p group, sh-circ_0023990+anti-miR-NC group, miR-873-5p group, miR-NC group, miR-873-5p+pcDNA-ANXA2 group and miR-873-5p+pcDNA group, and then clone formation experiment was used to detect cell radiosensitivity. After each group of cells was irradiated with 4Gy radiation, the expression of γH2AX protein in the cells was detected by Western Blot. The dual luciferase reporter gene experiment verified the targeting relationship between circ_0023990 and miR-873-5p or miR-873-5p and ANXA2. Results: The expression of circ_0023990 in thyroid cancer tissues was higher than that in normal tissues (2.15±0.09 vs. 0.97±0.05, P<0.05), and its expression was closely related to tumor size, lymph node metastasis and TNM staging of patients with thyroid cancer (P<0.05). The expression of circ_0023990 in thyroid cancer cell lines (TPC-1, KTC-1, FTC-133 and CAL-62) were higher than that of normal thyroid cells HTori-3 (3.16±0.38, 2.63±0.28, 1.82±0.24, 1.71±0.22 vs. 1.00±0.10, all P<0.05). The survival scores of TPC-1 and KTC-1 cells in the sh-circ_0023990 group were significantly lower than those in the sh-NC group (P<0.05), and the sensitization ratios were 2.482, 1.643; The survival scores of TPC-1 and KTC-1 cells in the sh-circ_0023990+anti-miR-873-5p group were higher than those in the sh-circ_0023990+anti-miR-NC group (P<0.05), and the sensitization ratios were 0.305, 0.441, respectively. The survival scores of TPC-1 and KTC-1 cells in the miR-873-5p group were lower than those in the miR-NC group (P<0.05), and the sensitization ratios were 2.044, 1.653 respectively. The survival scores of TPC-1 and KTC-1 cells in the miR-873-5p+pcDNA-ANXA2 group was higher than that in the miR-873-5p+pcDNA group (P<0.05), and the sensitization ratios were 0.496, 0.686, respectively. The expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+sh-circ_0023990 group were higher than that in the 4 Gy+sh-NC group (2.68±0.27 vs. 1.87±0.25, 2.46±0.19 vs. 1.77±0.14; all P<0.05), but the expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+sh-circ_0023990+anti-miR-873-5p group were lower than that in the 4 Gy+sh-circ_0023990+anti-miR-NC group (1.13±0.09 vs. 1.69±0.09, 1.11±0.08 vs. 1.60±0.08; both P<0.05). The expression of γH2AX protein in TPC-1 and KTC-1 cells in the 4 Gy+miR-873-5p group were higher than that in the 4 Gy+miR-NC group (2.35±0.16 vs. 1.84±0.14, 2.26±0.12 vs. 1.77±0.13; both P<0.05), but the expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+miR-873-5p+pcDNA-ANXA2 group were lower than that in the 4 Gy+miR-873-5p+pcDNA group (1.96±0.12 vs. 2.41±0.12, 1.92±0.07 vs. 2.28±0.12; both P<0.05). circ_0023990 targeted the negative regulation of miR-873-5p, and ANXA2 was the target gene of miR-873-5p. Conclusion: circ_0023990 was highly expressed in thyroid cancer tissues and cell lines, and it may promote the radiotherapy resistance of thyroid cancer cells in vivo through regulating miR-873-5p/ANXA2 axis.

First Evidence of Local E×B Drift in the Divertor Influencing the Structure and Stability of Confined Plasma near the Edge of Fusion Devices.

The structure of the edge plasma in a magnetic confinement system has a strong impact on the overall plasma performance. We uncover for the first time a magnetic-field-direction dependent density shelf, i.e., local flattening of the density radial profile near the magnetic separatrix, in high confinement plasmas with low edge collisionality in the DIII-D tokamak. The density shelf is correlated with a doubly peaked density profile near the divertor target plate, which tends to occur for operation with the ion B×∇B drift direction away from the X-point, as currently employed for DIII-D advanced tokamak scenarios. This double-peaked divertor plasma profile is connected via the E×B drifts, arising from a strong radial electric field induced by the radial electron temperature gradient near the divertor target. The drifts lead to the reversal of the poloidal flow above the divertor target, resulting in the formation of the density shelf. The edge density shelf can be further enhanced at higher heating power, preventing large, periodic bursts of the plasma, i.e., edge-localized modes, in the edge region, consistent with ideal magnetohydrodynamics calculations.

[Predictive value of vascular health indicators on newly cardiovascular events: Preliminary validation of Beijing vascular health stratification system].

OBJECTIVE: To explore the predictive value of carotid femoral artery pulse wave velocity (CF-PWV), carotid radial artery pulse wave velocity (CR-PWV), cardio-ankle vascular index (CAVI), and ankle brachial index (ABI) on coronary heart disease (CHD) and cerebral infarction (CI), and the preliminary validation of Beijing vascular health stratification (BVHS). METHODS: Subjects with at least 2 in-patient records were included into the study between 2010 and 2017 from Vascular Medicine Center of Peking University Shougang Hospital. Subjects with CHD or CI, and without data of vascular function at baseline were excluded. Eventually, 467 subjects free of CHD [cohort 1, mean age: (63.4±12.3) years, female 42.2%] and 658 subjects free of CI [cohort 2, mean age: (64.3±12.2) years, female 48.7%] at baseline were included. The first in-patient records were as the baseline data, the second in-patient records were as a following-up data. Cox proportional hazard regression was used to establish the predictive models of CHD or CI derived from BVHS by multivariable-adjusted analysis. RESULTS: The median follow-up time of cohort 1 and cohort 2 was 1.9 years and 2.1 years, respectively. During the follow-up, 164 first CHD events occurred in cohort 1 and 117 first CI events occurred in cohort 2. Four indicators were assessed as continuous variables simultaneously by multivariable-adjusted analysis. In cohort 1, CF-PWV, CR-PWV, ABI, and CAVI reached statistical significance in the multivariable-adjusted models (P<0.05). In cohort 2, only CAVI (P<0.05) was of statistical significance. In addition, the higher CF-PWV became a protector of CHD or CI (P<0.05). The prediction value of BVHS reached the statistical significance for CHD and CI in the unadjusted models (all P<0.05), however, BVHS could only predict the incidence of CHD (P<0.05), but not the incidence of CI (P>0.05) in the multivariable-adjusted models. CF-PWV, CR-PWV, ABI, and CAVI were associated factors of CHD independent of each other (P<0.05), only CAVI (P<0.05) was the risk factor of CI independent of the other three. CONCLUSION: The different vascular indicators might have different effect on CHD or CI. CAVI might be a stable predictor of both CHD and CI. Higher baseline CF-PWV was not necessarily a risk factor of CHD or CI because of proper vascular health management. BVHS was a potential factor for the prediction of CHD, and further research is needed to explore the prediction value for CI.

[Feasibility study of contrast injection scheme based on body mass and iodine flow rate in coronary computed tomography angiography with wide detector CT].

Objective: To investigate the feasibility of coronary computed tomography angiography (CCTA) with the individualized contrast medium injection scheme based on body mass and iodine flow rate by using wide detector CT at 100 kV. Methods: From May 2018 to July 2018,270 patients with clinically suspected coronary artery disease underwent CCTA in TEDA International Cardiovascular Hospital. They were randomly divided into three groups, 90 cases in each group: group A was 0.5 ml/kg, group B was 0.6 ml/kg, the flow rate of contrast medium injection in group A and group B was (body mass×25 mg)/the concentration of contrast medium,group C was 0.8 ml/kg and the flow rate of contrast medium was 5 ml/s. Iohexol (iodine concentration 350 mgI/ml) was used in all three groups. The CT values of the aorta root in the three groups, the CT value of the middle segment of LAD and RCA, SNR、CNR、SD of the aortic root, subjective score of coronary artery and effective dose (ED) were compared. Results: There was no significant difference in CT value of aortic root, CT value of middle LAD and RCA, SNR、CNR、SD of the aortic root, subjective score of coronary artery among group A, group B and group C (all P>0.05). The flow rates of the three groups of contrast agents were statistically different (4.5±0.5 vs 4.5±0.5 vs 5, P=0.015), and there was no statistical difference between group A and group B (P>0.05) but lower than group C (P=0.015); the amount of contrast agent in the three groups was statistically different (32±3 vs 38±4 vs 53±7, P<0.001), group A

Effects of absorbents on growth performance, blood profiles and liver gene expression in broilers fed diets naturally contaminated with aflatoxin.

OBJECTIVE: The study was conducted to evaluate the effects of the absorbent (a mixture of activated carbon and hydrated sodium calcium aluminosilicate) on growth performance, blood profiles and hepatic genes expression in broilers fed diets naturally contaminated with aflatoxin. METHODS: A total of 1200 one-day-old male chicks were randomly assigned to 6 treatments with 10 replicate cages per treatment. The dietary treatments were as follows: (1) control (basal diets); (2) 50% contaminated corn; (3) 100% contaminated corn; (4) control + 1% adsorbent; (5) 50% contaminated corn + 1% absorbent; (6) 100% contaminated corn + 1% absorbent. RESULTS: During d 1 to 21, feeding contaminated diets reduced (p&lt;0.05) body weight (BW), average daily gain (ADG) and average daily feed intake (ADFI), but increased (p&lt;0.05) feed-to-gain ratio (F/G). The absorbent supplementation increased (p&lt;0.05) BW, ADG and ADFI. There were interactions (p&lt;0.05) in BW, ADG and ADFI between contaminated corn and absorbent. Overall, birds fed 100% contaminated diets had lower (p&lt;0.05) final BW and ADG, but higher (p&lt;0.05) F/G compared to those fed control diets. The absorbent addition increased (p&lt;0.05) serum albumin concentration on d 14 and 28 and total protein (TP) level on d 28, decreased (p&lt;0.05) alanine transaminase activity on d 14 and activities of aspartate aminotransferase and alkaline phosphatase on d 28. Feeding contaminated diets reduced (p&lt;0.05) hepatic TP content on d 28 and 42. The contaminated diets upregulated (p&lt;0.05) expression of interleukin-6, catalase (CAT) and superoxide dismutase (SOD), but downregulated (p&lt;0.05) glutathione S-transferase (GST) expression in liver. The absorbent supplementation increased (p&lt;0.05) interleukin-1ß, CAT, SOD, cytochrome P450 1A1 and GST expression in liver. There were interactions (p&lt;0.05) in the expression of hepatic CAT, SOD and GST between contaminated corn and absorbent. CONCLUSION: The results suggest that the naturally aflatoxin-contaminated corn depressed growth performance, while the adsorbent could partially attenuate the adverse effects of aflatoxin on growth performance, blood profiles and hepatic genes expression in broilers.

Effects of stale maize on growth performance, immunity, intestinal morphology and antioxidant capacity in broilers.

OBJECTIVE: This study was conducted to determine the effects of stale maize on growth performance, immunity, intestinal morphology, and antioxidant capacity in broilers. METHODS: A total of 800 one-day-old male Arbor Acres broilers (45.4±0.5 g) were blocked based on body weight, and then allocated randomly to 2 treatments with 20 cages per treatment and 20 broilers per cage in this 6-week experiment. Dietary treatments included a basal diet and diets with 100% of control maize replaced by stale maize. RESULTS: The content of fat acidity value was higher (p<0.05) while the starch, activities of catalase and peroxidase were lower (p<0.05) than the control maize. Feeding stale maize diets reduced (p<0.05) average daily feed intake (ADFI) throughout the experiment, feed conversion ratio (FCR) during d 0 to 21 and the whole experiment as well as relative weight of liver, spleen, bursa of Fabricius and thymus (p<0.05) on d 21. Feeding stale maize diets decreased jejunum villus height (VH) and VH/crypt depth (CD) (p<0.05) on d 21 and 42 as well as ileum VH/CD on d 42. The levels of immunoglobulin G, acid α-naphthylacetate esterase positive ratios and lymphocyte proliferation on d 21 and 42 as well as lysozyme activity and avian influenza antibody H5N1 titer on d 21 decreased (p<0.05) by the stale maize. Feeding stale maize diets reduced (p<0.05) serum interferon-γ, tumor necrosis factor-α, interleukin-2 on d 21 and interleukin-6 on d 21 and 42. Broilers fed stale maize diets had lower levels of (p<0.05) total antioxidative capacity on d 42, superoxide dismutase and glutathione peroxidase on d 21 and 42, but higher (p<0.05) levels of malondialdehyde on d 21 and 42. CONCLUSION: Feeding 100% stale maize decreased ADFI and FCR, caused adverse effects on immunity and antioxidant function and altered intestinal morphology in broilers.

Effect of probiotics and xylo-oligosaccharide supplementation on nutrient digestibility, intestinal health and noxious gas emission in weanling pigs.

OBJECTIVE: This study was conducted to evaluate the effect of probiotics (Bacillus subtilis and Enterococcus faecium) and xylo-oligosaccharide (XOS) supplementation on growth performance, nutrient digestibility, serum profiles, intestinal health, fecal microbiota and noxious gas emission in weanling pigs. METHODS: A total of 240 weanling pigs ([Yorkshire×Landrace]×Duroc) with an average body weight (BW) of 6.3±0.15 kg were used in this 28-day trial. Pigs were randomly allocated in 1 of the following 4 dietary treatments in a 2×2 factorial arrangement with 2 levels of probiotics (0 and 500 mg/kg probiotics) and XOS (0 and 200 mg/kg XOS) based on the BW and sex. RESULTS: Administration of probiotics or XOS improved average daily gain (p<0.05) during 0 to 14 d and the overall period, while pigs that were treated with XOS had a greater average daily gain and feed efficiency (p<0.05) compared with unsupplemented treatments throughout 15 to 28 d and the whole experiment. Either probiotics or XOS treatments increased the apparent total tract digestibility of nutrients (p<0.05) during 0 to 14 d. No effects on serum profiles were observed among treatments. The XOS increased villus height: crypt depth ratio in jejunum (p<0.05). The supplementation of probiotics (500 mg/kg) or XOS (200 mg/kg) alone improved the apparent total tract digestibility of dry matter, nitrogen and gross energy on d 14, the activity of trypsin and decreased fecal NH3 concentration (p<0.05). Administration of XOS decreased fecal Escherichia coli counts (p<0.05), while increased lactobacilli (p<0.05) on d 14. There was no interaction between dietary supplementation of probiotics and XOS. CONCLUSION: Inclusion of XOS at 200 mg/kg or probiotics (Bacillus subtilis and Enterococcus faecium) at 500 mg/kg in diets containing no antibiotics significantly improved the growth performance of weanling pigs. Once XOS is supplemented, further providing of probiotics is not needed since it exerts little additional effects.

Effects of dietary energy and lipase levels on nutrient digestibility, digestive physiology and noxious gas emission in weaning pigs.

OBJECTIVE: This study was conducted to evaluate the effect of dietary energy and lipase supplementation on growth performance, nutrient digestibility, serum profiles, intestinal morphology, small intestinal digestive enzyme activities, biochemical index of intestinal development and noxious gas emission in weaning pigs. METHODS: A total of 240 weaning pigs ([Yorkshire×Landrace]×Duroc) with an average body weight (BW) of 7.3±0.12 kg were used in this 28-d experiment. Weaning pigs were randomly allocated to 4 dietary treatments in a 2×2 factorial arrangement with 2 levels of energy (net energy = 2,470 kcal/kg for low energy diet and 2,545 kcal/kg for basal diet) and 2 levels of lipase (0 and 1.5 U/g of lipase) according to BW and sex. There were 6 replications (pens) per treatment and 10 pigs per pen (5 barrows and 5 gilts). RESULTS: Weaning pigs fed the low energy diet had lower (p<0.05) gain-to-feed ratio (G:F) throughout the experiment, apparent digestibility of dry matter, nitrogen, ether extract, and gross energy during d 0 to 14, average daily gain during d 15 to 28, lipase activity in duodenum and ileum and protein/DNA in jejunum (p<0.05), respectively. Lipase supplementation had no effect on growth performance but affected apparent nutrient digestibility (p<0.05) on d 14 and enhanced lipase activity in the duodenum and ileum and protease activity in duodenum and jejunum of pigs (p<0.05) fed the low energy diet. Lipase reduced serum low-density lipoprotein cholesterol (LDL-C) and triglyceride (TG), NH3 production (p<0.05) from the feces. CONCLUSION: The low energy diet decreased G:F throughout the experiment and nutrient digestibility during d 0 to 14 as well as lipase activity in duodenum and ileum. Lipase supplementation increased nutrient digestibility during d 0 to 14 and exerted beneficial effects on lipase activity in duodenum and ileum as well as protease activity in duodenum and jejunum, while reduced serum LDL-C, TG and fecal NH3.

MiR-200c inhibits metastasis of breast tumor via the downregulation of Foxf2.

The forkhead box F2 (Foxf2) gene suppresses epithelial-mesenchymal transition via the modulation of transcription of zinc finger E-box-binding homeobox 1 (Zeb1) and epithelial (E)-cadherin, thereby inhibiting tumor metastasis. Additionally, the specific binding of microRNA (miR)-200c to Foxf2 mRNA impedes metastatic pulmonary cancer. However, the role of miR-200c in breast cancer is still unknown. Therefore, in this study, miR-200c mimics were transfected into the highly metastatic breast cancer cell line MDA-MB-231. Their invasion and migration abilities were observed by scratch and transwell migration assays. Real-time PCR was used to detect mRNA levels of Foxf2, Zeb1, and E-cadherin, whereas Foxf2 protein level was determined by western blot analysis. Our results showed that, compared to the control group, miR-200c inhibited the migration or invasion of MDA-MB-231 cells. Real-time PCR and western blot analysis exhibited significant decreases in Foxf2 expression in the presence of miR-200c, along with a decrease in Zeb1 and increase in E-cadherin mRNA expressions. Thus, our preliminary data demonstrated that miR-200c could inhibit the metastasis of breast cancer cells by downregulating Foxf2 expression, providing leads for the discovery of a novel breast cancer treatment.

Effects of feeding frequency on meat quality traits and Longissimus muscle proteome in finishing pigs.

Sixty-four barrows with an initial body weight of 59.8 ± 2.1 kg were allocated to one of the two feeding frequency regimes (had free access to diet and fed two meals per day). Pigs had free access (FA) to feed were fed on an ad libitum basis during the 8-week experimental period. Pigs fed twice daily (M2) were allowed to consume their meals in 2 h. Pigs fed twice daily had lower average daily feed intake (p < 0.01) and average daily gain (p < 0.1), but a greater G:F (p < 0.05) than FA pigs. Lower perirenal fat deposition, hot carcass weight, intramuscular fat content (p < 0.05) and dressing percentage (p < 0.1) were found in M2 pigs compared with FA pigs. Activities of citrate synthase, ß-hydroxylacyl-CoA dehydrogenase and lactate dehydrogenase were greater in the Longissimus muscle (LM) of M2 pigs compared with FA pigs (p < 0.05). Proteomic analysis revealed that expression abundances of proteins involved in glucose metabolism, energy production and lipid utilization were upregulated, but expression levels of proteins participating in protein and amino acid metabolism, stress response and redox homeostasis were downregulated in the LM of M2 pigs than those in FA pigs (p < 0.05). In conclusion, the less meal frequency impairs growth rate, has marginal effects on carcass and meat quality traits and affects expression abundances of proteins in the LM of finishing pigs.

[Relationship between large artery elasticity function and coronary heart disease, lower extremity arterial disease in patients with carotid plaque].

OBJECTIVE: To evaluate the relationship between large artery elastic function and coronary heart disease (CHD) or lower extremity arterial disease (LEAD) in patients with carotid plaque. METHODS: A total of 491 patients with carotid plaque were enrolled into the study with complete data of arterial stiffness detection and blood test [male: 208 and female: 283, and mean age: (61.66±11.60) years]. All the subjects were divided into 2 groups according to CHD or LEAD, namely non-CHD&LEAD group (neither CHD nor LEAD) and CHD/LEAD group (either CHD or LEAD). Accor-ding to the mean age level (age<61.66 years or age>61.66 years), the independent association was analyzed between higher large arterial stiffness (carotid-femoral pulse wave velocity, CF-PWV, CF-PWV>9 m/s) and CHD/LEAD. RESULTS: In the present research population, the mean level of arterial stiff-ness was high (the mean CF-PWV was 10.71 m/s), and 76.6% of them had arteriosclerosis, and 36.9% CHD/LEAD. The age, male and smoking proportion, systolic blood pressure (SBP), glycosylated hemoglobin (HbA1c), homocysteine (Hcy), creatinine (Cr), CF-PWV, prevalence rate of hypertension and diabetes mellitus, medication on hypertension, diabetes and hyperlipidemia were higher in CHD/LEAD group, and total cholesterol (TC), high density lipoprotein cholesterol (HDL-C), and low density lipoprotein cholesterol (LDL-C) were lower in CHD/LEAD group than in non-CHD&LEAD group (all P<0.05).In multivariate Logistic regression analysis, the results showed that in the patients with age below 61.66 years, large artery stiffness (CF-PWV>9 m/s) was an independent risk factor of CHD/LEAD (OR=3.229, 95%CI 1.156-9.022, P<0.05); In the patients with age above 61.66 years, there was no independent association between large artery stiffness and CHD/LEAD (P>0.05). CONCLUSION: The large artery elasticity function in the patients with carotid plaque was poor. In the patients with carotid plaque and higher large artery stiffness below 61.66 years, the risk of the prevalence of CHD/LEAD was increased significantly than with normal arterial stiffness. In the patients with carotid plaque below or above 61.66 years, the independent influencing factors on the prevalence of CHD/LEAD were different.

Casein Supplementation Does Not Affect the Estimates of True Total Tract Digestibility of Phosphorus in Soybean Meal for Growing Pigs Determined by the Regression Method.

Forty-eight barrows with an average initial body weight of 25.5±0.3 kg were assigned to 6 dietary treatments arranged in a 3×2 factorial of 3 graded levels of P at 1.42, 2.07, or 2.72 g/kg, and 2 levels of casein at 0 or 50 g/kg to compare the estimates of true total tract digestibility (TTTD) of P in soybean meal (SBM) for pigs fed diets with or without casein supplementation. The SBM is the only source of P in diets without casein, and in the diet with added casein, 1.0 to 2.4 g/kg of total dietary P was supplied by SBM as dietary level of SBM increased. The experiment consisted of a 5-d adjustment period and a 5-d total collection period with ferric oxide as a maker to indicate the initiation and termination of fecal collection. There were interactive effects of casein supplementation and total dietary P level on the apparent total tract digestibility (ATTD) and retention of P (p<0.05). Dietary P intake, fecal P output, digested P and retained P were increased linearly with graded increasing levels of SBM in diets regardless of casein addition (p<0.01). Compared with diets without casein, there was a reduction in fecal P in the casein-supplemented diets, which led to increases in digested P, retained P, ATTD, and retention of P (p<0.01). Digested N, ATTD of N, retained N, and N retention were affected by the interaction of casein supplementation and dietary P level (p<0.05). Fecal N output, urinary N output, digested N, and retained N increased linearly with graded increasing levels of SBM for each type of diet (p<0.01). The estimates of TTTD of P in SBM, derived from the regression of daily digested P against daily P intake, for pigs fed diets without casein and with casein were calculated to be 37.3% and 38.6%, respectively. Regressing daily digested N against daily N intake, the TTTD of N in SBM were determined at 94.3% and 94.4% for diets without casein and with added casein, respectively. There was no difference in determined values of TTTD of P or N in SBM for pigs fed diets with or without casein (p>0.05). In summary, our results demonstrate that the estimates of TTTD of P in SBM for pigs were not affected by constant casein inclusion in the basal diets.

Influence of colorectal cancer tumor suppressor gene CHD5 methylation on its clinical and pathological characteristics.

Recently, abnormal tumor suppressor gene (TSG) methylation has become a hotspot in the research on colorectal cancer (CRC). This study aimed to explore the influence of CHD5 methylation of CRC TSG on its clinical and pathological characteristics. A total of 40 operation samples as well as corresponding tissue specimens were collected from CRC patients treated in the First Affiliated Hospital of Zhengzhou University from January to December in 2014. CHD5 gene methylation in tissue specimens was detected with methylation specific polymerase chain reaction (MSP); moreover, messenger ribose nucleic acid (mRNA) expression of CHD5 in each tissue was tested using reverse transcription-polymerase chain reaction (RT-PCR), and Western blot was applied to detect the expression of CHD5 protein in those tissues and to analyze the correlation between mRNA and protein of cancer tissue CHD5 as well as the relationship between CHD5 methylation and protein expression. Results revealed that the expression rate of CHD5 methylation in 40 normal mucosal tissues, para-carcinoma tissues, adenoma tissues and CRC tissues was 12.5% (5/40), 22.5% (9/40), 47.5% (19/40) and 72.5% (33/40), respectively. The mRNA expression of CHD5 in the above tissues was 0.225±0.276, 0.169±0.231, 0.147±0.159 and 0.013±0.011 and the protein expression of CHD5 was 0.438±0.205, 0.398±0.180, 0.156±0.1 and 0.024±0.311, respectively. Methylation rate of CHD5 was 87% (20/23) in 23 cases of CHD5 protein loss expression and 52.9% (9/17) in 17 cases of CHD5 protein expression. Results of chi-squared test indicated that there was a significant difference in methylation rate (P less than 0.05), that is, the methylation rate of negatively expressed CHD5 protein was obviously higher than positively expressed protein. Thus, it can be concluded that the CHD5 methylation rate rises gradually in the evolution of CRC, which is related to the occurrence and development of CRC. Furthermore, CHD5 mRNA is positively correlated with protein expression and CHD5 gene methylation is associated with protein loss expression. Therefore, TSG CHD5 methylation of rectal cancer has a great effect in influencing its clinical and pathological features.

Migrations of pentagon-heptagon defects in hexagonal boron nitride monolayer: the first-principles study.

The first-principles calculations are employed to study the migrations of pentagon-heptagon (5-7) defects in hexagonal boron nitride monolayer (h-BN). A type of grain boundaries, consisted of 5-7 defects, is constructed on the basis of experimental observations. With the absorption of a pair of atoms, one 5-7 defect in the grain boundary migrates apart by one unit cell and afterward migrates again through the bond rotation. It is also found that the two migrations could be replaced by one single step when the pair of absorbed atoms is located at another specific site in the same heptagon. Energy barriers and reaction paths for the migrations of 5-7 defects in h-BN by the bond rotation are theoretically investigated by the standard nudged elastic band method and the generalized solid-state nudged elastic band method. To elucidate the difference between the bond rotation process of the 5-7 defects with N-N bonds and those with B-B bonds, a couple of typical 21.7° grain boundaries with either N-N or B-B bonds are investigated. It is shown that the energy barrier of the migration of defects with N-N bonds is lower than that with B-B bonds in this type of grain boundaries.

Multivariate analysis of molecular markers in peripheral blood associated with recurrence and metastasis of hepatocellular carcinoma.

Invasion, metastasis, and recurrence are the most common causes of death in patients with hepatocellular carcinoma (HCC) and are therefore critical factors for both therapy and prognosis. Current methods for diagnosis of HCC rely mainly on serological markers such as alpha-fetoprotein and liver enzymes, together with physical assessment and imaging techniques. The availability of more accurate serum markers may facilitate screening and early diagnosis, which will improve prognosis. This retrospective cohort analysis included 50 consecutive patients with cirrhosis and single or multifocal HCC and 40 control subjects with no liver disease or risk factors for viral hepatitis. Expression of epidermal growth factor-like domain 7 (EGFL7), osteopontin (OPN), and prostaglandin E2 (PGE2) were detected using an enzyme-linked immunosorbent assay. The mean serum levels of EGFL7, OPN, and PGE2 in the HCC group were 132.11 pg/mL, 11.77 ng/mL, and 179.37 pg/mL, respectively, which were all significantly higher than the levels in the control group (23.03 pg/mL, 2.31 ng/mL, and 47.36 pg/mL, respectively; P < 0.001). Serum levels of EGFL7, OPN, and PGE2 levels may thus be useful for screening and surveillance of HCC among high-risk populations, and have the potential to improve prognosis of these patients.

Analysis of agouti signaling protein (ASIP) gene polymorphisms and association with coat color in Tibetan sheep (Ovis aries).

Tibetan sheep, an indigenous breed, have a wide variety of phenotypes and a colorful coat, which make this breed an interesting model for evaluating the effects of coat-color gene mutations on this phenotypic trait. The agouti signaling protein (ASIP) gene is a positional candidate gene, as was inferred based on previous study. In our research, ASIP gene copy numbers in genomic DNA were detected using a novel approach, and the exon 2 g.100-104 mutation and copy number variation (CNV) of ASIP were associated with coat color in 256 sheep collected from eight populations with different coat colors by high-resolution melting curve assay. We found that the relative copy numbers of ASIP ranged from one to eight in Tibetan sheep. All of the g.100-104 genotypes in the populations were in Hardy-Weinberg equilibrium, and there was no relationship between the g.100-104 genotype and coat color (P > 0.05). The single ASIP CNV allele was found to be almost entirely associated with solid-black coat color; however, not all solid-black sheep displayed the putative single ASIP CNV genotype. From our study, we speculate that the ASIP CNV is under great selective pressure and the single ASIP CNV allows selection for black coat color in Tibetan sheep, but this does not explain all black phenotypes in Tibetan sheep.

De novo assembly and characterization of skin transcriptome using RNAseq in sheep (Ovis aries).

Wool is produced via synthetic processes of wool follicles, which are embedded in the skin of sheep. The development of new-generation sequencing and RNA sequencing provides new approaches that may elucidate the molecular regulation mechanism of wool follicle development and facilitate enhanced selection for wool traits through gene-assisted selection or targeted gene manipulation. We performed de novo transcriptome sequencing of skin using the Illumina Hiseq 2000 sequencing system in sheep (Ovis aries). Transcriptome de novo assembly was carried out via short-read assembly programs, including SOAPdenovo and ESTScan. The protein function, clusters of orthologous group function, gene ontology function, metabolic pathway analysis, and protein coding region prediction of unigenes were annotated by BLASTx, BLAST2GO, and ESTScan. More than 26,266,670 clean reads were collected and assembled into 79,741 unigene sequences, with a final assembly length of 35,447,962 nucleotides. A total of 22,164 unigenes were annotated, accounting for 36.27% of the total number of unigenes, which were divided into 25 classes belonging to 218 signaling pathways. Among them, there were 17 signal paths related to hair follicle development. Based on mass sequencing data of sheepskin obtained by RNA-Seq, many unigenes were identified and annotated, which provides an excellent platform for future sheep genetic and functional genomic research. The data could be used for improving wool quality and as a model for human hair follicle development or disease prevention.

The effect of antifibrotic drug halofugine on Th17 cells in concanavalin A-induced liver fibrosis.

Anti-inflammation strategy is one of the proposed therapeutic approaches to hepatic fibrosis. T helper (Th) 17 cells, which play a detrimental role in experimental murine models of inflammatory diseases, have been demonstrated to participate in the pathogenesis of liver damage. The inhibitory effect of halofuginone (HF), an active component of extracts derived from the plant alkaloid febrifugine, on collagen synthesis has been shown in animal models of the fibrotic disease. The aim of this study was to clarify the in vivo effect of HF on Th17 cells in concanavalin A-induced fibrosis rats. Haematoxylin-eosin (HE) staining and Masson staining were performed to observe collagen deposition. The presence of INF-gamma, TNF-alpha, IL-6, IL-17, IL-1beta, IL-33 and IL-10 in serum and the presence of ROR-γt, IL-17, TGF-ß1 and α-SMA in liver tissue were detected. Flow cytometry was performed to analyse the percentage of Th17 cells. We observed significantly lower levels of INF-gamma, TNF-alpha, IL-6, IL-17, IL-1beta, TGF-ß1 and α-SMA in HF-treated group of rats, and the percentage of Th17 cells in splenic lymphocyte was decreased well. Histological examination demonstrated that HF significantly reduced the severity of liver fibrosis in HF-treated rats. We concluded that HF (10 mg/kg) exerts an antifibrotic impact on Th17 cells and its relative cytokines in rats with ConA-induced fibrosis.