[Advances in the classification and treatment of isolated superior mesenteric artery dissection].

Isolated superior mesenteric artery dissection (ISMAD) has attracted more and more clinicians' attention in recent years. Patients onset of ISMAD often present with abdominal pain. The misdiagnosis or miss diagnosis is common because of the non-specific symptoms and signs, which even can endanger lives in serious cases. Imaging classification is of great significance for diagnosis and treatment of ISMAD. The Sakamoto classification and the Yun classification are two classical classified methods. However, with the further study of ISMAD, various new classifications emerge. Conservative treatment was once considered as the preferred. As the rapid development of endovascular therapy and the great progress of new devices, stenting therapy can significantly improve symptoms and achieve satisfactory long-term effects, and be even expected to become the preferred method for clinical therapy of ISMAD. However, the long-term effects of endovascular therapy still need a large number of follow-up data, and complications after stent implantation can't be ignored.

Fetal growth at different gestational periods and risk of impaired childhood growth, low childhood weight and obesity: a prospective birth cohort study.

OBJECTIVE: To examine the longitudinal associations of fetal growth with adverse child growth outcomes and to assess whether maternal metabolic factors modify the associations. DESIGN: Prospective cohort study. SETTING: Born in Guangzhou Cohort Study, China. POPULATION: A total of 4818 mother-child pairs. METHODS: Fetal growth was assessed according to estimated fetal weight (EFW) from 22 weeks of gestation until birth and the measurement of the birthweight. Fetal growth Z-scores were computed from random effects in the multilevel linear spline models to represent fetal size in early pregnancy (22 weeks of gestation) and growth in mid-pregnancy (22-27 weeks of gestation), early third trimester (28-36 weeks of gestation) and late third trimester (≥37 weeks of gestation). MAIN OUTCOME MEASURES: Z-scores for childhood stunting, low weight, overweight or obesity, length/height for age (LAZ/HAZ), weight for age (WAZ) and body mass index for age (BMIZ) at the age of 3 years. Adjusted associations were examined using multiple Poisson or linear regression models. RESULTS: Increased Z-scores of fetal size in early pregnancy and growth in mid-pregnancy and early third trimester were associated with a higher risk of childhood overweight or obesity (risk ratios 1.25-1.45). Fetal growth in each period was negatively associated with stunting and low weight, with the strongest associations observed for fetal size in early pregnancy and growth in mid-pregnancy. The results for continuous outcomes (LAZ/HAZ, WAZ and BMIZ) were similar. The associations of fetal growth with overweight or obesity in childhood were stronger among mothers who were underweight and who were overweight or obese than among mothers of normal weight. CONCLUSIONS: Accelerated fetal growth before 37 weeks of gestation is associated with children who are overweight or obese, whereas the critical period for stunting and low weight occurs before 28 weeks of gestation. TWEETABLE ABSTRACT: Fetal growth during different periods is differentially associated with childhood stunting, underweight and overweight or obesity.

[A Luciferase-EGFP Reporter System for the Evaluation of DNA Methylation in Mammalian Cells].

DNA methylation is an essential epigenetic modification involved in numerous biological processes. Here, we present a cell-based system pLTR-Luc2P-EGFP for evaluation of DNA methylation in mammalian cells. In this system, the expression of reporter gene luciferase2P (Luc2P)-EGFP is under the control of HIV-1 promoter 5' long terminal repeat (LTR), which contains multiple CpG sites. Once these sites are methylated, the expression of Luc2P-EGFP is turned off, which may be visualized under fluorescence microscopy, with quantification performed in luciferase activity assay. As a proof of principle, pLTR-Luc2P-EGFP was methylated in vitro, and transfected into 293T cells, where the reduction of Luc2P-EGFP expression was confirmed. Premixed reporter DNA samples with the methylation levels varying from 0 to 100% were used for quantitative measurements of DNA methylation. The resulting standard curves indicated the accuracy of luciferase activity exceeding that of the Western blotting against EGFP. The Bland-Altman analysis showed that data from luciferase activity assay were in good agreement with the actual DNA methylation levels. In summary, we have established a reporter system coupled with reliable detection technique capable of efficient quantifying the changes in methylation in mammalian cells. This system may be utilized as a high throughput screening tool for identifying molecules that modulate DNA methylation.

[The status and related factors of myopia for children and adolescents aged 5-18 years old in Shaanxi Province in 2018].

In 2018, the myopia detection rate of children and adolescents aged 5-18 years old in Shaanxi Province was 54.9% (11 060/20 144). The myopia detection rate of girls ï¼»58.6%(5 830/9 949)ï¼½ was higher than that of boys ï¼»58.6% (3 416/5 830)ï¼½ (P<0.001). In children and adolescents, the myopia detection rate increased with the age before their 16 years old, and saw a stable or downward trend after the age of 16. After adjusting the confounding factors, the myopia detection rate of children and adolescents from regions with per capita GDP>100 000 yuan was higherthan that of children and adolescents from regions with per capita GDP<50 000 yuanï¼»OR (95%CI):1.58(1.34-1.87)ï¼½.

Safety, efficacy and tumor mutational burden as a biomarker of overall survival benefit in chemo-refractory gastric cancer treated with toripalimab, a PD-1 antibody in phase Ib/II clinical trial NCT02915432.

BACKGROUND: High tumor mutational burden (TMB-H) is correlated with enhanced objective response rate (ORR) and progression-free survival (PFS) for certain cancers receiving immunotherapy. This study aimed to investigate the safety and efficacy of toripalimab, a humanized programmed death-1 (PD-1) antibody, in advanced gastric cancer (AGC), and the predictive survival benefit of TMB and PD-L1. PATIENTS AND METHODS: We reported on the AGC cohort of phase Ib/II trial evaluating the safety and activity of toripalimab in patients with AGC, oesophageal squamous cell carcinoma, nasopharyngeal carcinoma and head and neck squamous cell carcinoma. In cohort 1, 58 chemo-refractory AGC patients received toripalimab (3 mg/kg d1, Q2W) as a monotherapy. In cohort 2, 18 chemotherapy-naive AGC patients received toripalimab (360 mg d1, Q3W) with oxaliplatin 130 mg/m2 qd, d1, capecitabine 1000 mg/m2 b.i.d., d1-d14, Q3W as first-line treatment. Primary end point was ORR. Biomarkers such as PD-L1 and TMB were evaluated for correlation with clinical efficacy. RESULTS: In cohort 1, the ORR was 12.1% and the disease control rate (DCR) was 39.7%. Median PFS was 1.9 months and median OS was 4.8 months. The TMB-H group showed significant superior OS than the TMB-L group [14.6 versus 4.0 months, HR = 0.48 (96% CI 0.24-0.96), P = 0.038], while PD-L1 overexpression did not correlate with significant survival benefit. A 77.6% of patients experienced at least one treatment-related adverse event (TRAE), and 22.4% of patients experienced a grade 3 or higher TRAE. In cohort 2, the ORR was 66.7% and the DCR was 88.9%. A 94.4% of patients experienced at least one TRAE and 38.9% of patients experienced grade 3 or higher TRAEs. CONCLUSIONS: Toripalimab has demonstrated a manageable safety profile and promising antitumor activity in AGC patients, especially in combination with XELOX. High TMB may be a predictive marker for OS of AGC patients receiving toripalimab as a single agent. TRIAL REGISTRATION: ClinicalTrials.gov NCT02915432.

[Association between TV watching time and the cognitive functions among the elderly aged 60 years and above in Xiamen City].

Objective: To explore the cognitive functions of the elderly aged 60 years and above in Xiamen, and whether TV watching time would affect those functions. Methods: We conducted a cross-sectional study on 3 230 registered individuals aged 60 years and above in Xiamen from July to October in 2016 with a multi-stage random sampling method. Montreal Cognitive Assessment (MoCA) was used to measure cognitive functions and ordinal logistic regression was used to quantify their associations with TV watching time. Results: A total of 2 944 respondents were included in this study. The overall age of them was (70.14±7.81) years. 51.49% (n=1 516) of all participants were men. Overall, the mean scores of MoCA in general and each subdomain cognitive function were 18.65±6.43 (general), 5.43±1.17 (orientation), 2.33±1.95 (memory), 1.80±1.52 (visuospatial), 1.66±1.20 (executive), 4.54±1.71 (attention), and 4.09±1.64 (language). Compared to those who watched TV no more than 2 hours per day, the elderly who did not watch TV had a worse performance in general, orientation, memory, visuospatial, executive, attention, language function, with OR (95%CI) values about 0.50 (0.35-0.71), 0.52 (0.37-0.73), 0.68 (0.48-0.96), 0.67 (0.47-0.96), 0.48 (0.34-0.67), 0.56 (0.41-0.77) and 0.45 (0.33-0.62), respectively. As to respondents who watched TV more than 2 hours but no more than 4 hours per day, they had a better performance in general, executive, attention, language, and orientation function with respective OR (95%CI) values about 1.24 (1.05-1.47), 1.20 (1.02-1.41), 1.41 (1.19-1.67), 1.51 (1.28-1.77) and 1.33 (1.08-1.64). Conclusion: The elderly aged 60 years and above in Xiamen City with TV watching time between 2 and 4 hours per day had a good performance in cognitive functions compared to those with more or less TV watching time.

Recognition of Y Fragment Deletion by Genotyping Graphs after Amplified by PowerPlex® 21 Detection Kit.

OBJECTIVES: To recognize the possibility of Y fragment deletion of Amelogenin gene intuitively and simply according to the genotyping graphs. METHODS: By calculating the ratio of total peak height of genotyping graphs, the statistics of equilibrium distribution between Amelogenin and D3S1358 loci, Amelogenin X-gene and Amelogenin Y-gene, and different alleles of D3S1358 loci from 1 968 individuals was analyzed after amplified by PowerPlex® 21 detection kit. RESULTS: Sum of peak height of Amelogenin X allele was not less than 60% that of D3S1358 loci alleles in 90.8% female samples, and sum of peak height of Amelogenin X allele was not higher than 70% that of D3S1358 loci alleles in 94.9% male samples. CONCLUSIONS: The result of genotyping after amplified by PowerPlex® 21 detection kit shows that the possibility of Y fragment deletion should be considered when only Amelogenin X-gene of Amelogenin is detected and the peak height of Amelogenin X-gene is not higher than 70% of the total peak height of D3S1358 loci.

Hepatitis B virus infection is associated with gastric cancer in China: an endemic area of both diseases.

BACKGROUND: Hepatitis B virus (HBV) infection was demonstrated to be a risk factor of several cancers of the digestive system. In addition, liver cirrhosis, which could possibly result from chronic HBV infection, was associated with a higher risk of gastric cancer. However, the association of HBV infection and gastric cancer has not been investigated. METHODS: A retrospective case-control study with 580 cases and 580 controls matched for age, sex and year of diagnosis was conducted. The associations between gastric cancer and HBV infection were explored with univariate and multivariate unconditional logistic regression analysis. RESULTS: Hepatitis B surface antigen (HBsAg) was positively associated with gastric cancer (AOR (95% CI): 1.49 (1.06-2.10)). This association remained significant in patients without family history of gastric cancer (AOR (95% CI): (1.06-2.11)). For HBsAg-negative population, being anti-HBc positive/anti-HBs negative, which possibly indicated occult HBV infection, was also found to have some associations with gastric cancer. In addition, some synergistic effects between HBV infection and blood type A in gastric cancer were identified. CONCLUSIONS: The HBV infection was positively related with gastric cancer, especially for patients without family history of gastric cancer. Further prospective studies are warranted to confirm this relationship.

Transversally and axially tunable carbon nanotube resonators in situ fabricated and studied inside a scanning electron microscope.

We report a new design of carbon nanotube (CNT) resonator, whose resonance frequency can be tuned not only transversally by a gate voltage, but also by the axial strain applied through directly pulling the CNT. The resonators are fabricated from individual suspended single-walled CNT (SWCNT) in situ inside a scanning electron microscope. The resonance frequency of a SWCNT resonator can be tuned by more than 20 times with an increase of quality factor when the axial strain of the SWCNT is only increased from nearly zero to 2% at room temperature. The transversal gate-tuning ability is found to be weaker than the axial-tuning ability and decrease with increasing the axial strain. The gate voltage can hardly tune the resonance frequency when the initial axial strain is larger than 0.35% and the CNT acts like a tied string. The relationship among resonance frequency, gate voltage, and initial axial strain of the CNT obtained presently will allow for the designs of CNT resonators with high frequency and large tuning range. The present resonator also shows ultrahigh sensitivity in displacement and force detection, with a resolution being better than 2.4 pm and 0.55 pN, respectively.

Effects of exercise training on carbohydrate and lipid catabolism in the swimming muscles of Nile tilapia (Oreochromis niloticus).

This study aims to determine the effects of exercise training on carbohydrate and lipid catabolism in the swimming muscles of Nile tilapia (Oreochromis niloticus) by measuring the levels of related enzymes, lipids and free fatty acids. We designed one control group and two training groups of fish that were exercised at different training intensities [0, 1 and 1.5 body lengths per second (bl/s)]. The fish in the experimental groups were trained for 12 h/day for 4 weeks. Compared with the control group, the 1 and 1.5 bl/s groups showed significantly increased hexokinase and pyruvate kinase activities in red muscle (p < 0.05). In white muscle, pyruvate kinase activity was significantly higher in the 1.5 bl/s group than in the control group (p < 0.05), and hexokinase activity did not significantly differ between the groups. The activities of hormone-sensitive lipase and carnitine palmitoyltransferase I in both muscle types were significantly lower in the training groups than in the control group (p < 0.05). The plasma-free fatty acid level decreased (p < 0.05), while the lipid percentages increased in red muscle (p < 0.05) after exercise training. These findings clearly indicated that with exercise training, glycolysis increased and lipid oxidation decreased in the swimming muscle of tilapia.

In situ multiproperty measurements of individual nanomaterials in SEM and correlation with their atomic structures.

The relationship between property and structure is one of the most important fundamental questions in the field of nanomaterials and nanodevices. Understanding the multiproperties of a given nano-object also aids in the development of novel nanomaterials and nanodevices. In this paper, we develop for the first time a comprehensive platform for in situ multiproperty measurements of individual nanomaterials using a scanning electron microscope (SEM). Mechanical, electrical, electromechanical, optical, and photoelectronic properties of individual nanomaterials, with lengths that range from less than 200 nm to 20 µm, can be measured in situ with an SEM on the platform under precisely controlled single-axial strain and environment. An individual single-walled carbon nanotube (SWCNT) was measured on the platform. Three-terminal electronic measurements in a field effect transistor structure showed that the SWCNT was semiconducting and agreed with the structure characterization by transmission electron microscopy after the in situ measurements. Importantly, we observed a bandgap increase of this SWCNT with increasing axial strain, and for the first time, the experimental results quantitatively agree with theoretical predictions calculated using the chirality of the SWCNT. The vibration performance of the SWCNT, a double-walled CNT, and a triple-walled CNT were also studied as a function of axial strain, and were proved to be in good agreement with classical beam theory, although the CNTs only have one, two, or three atomic layers, respectively. Our platform has wide applications in correlating multiproperties of the same individual nanostructures with their atomic structures.

[Epidemic characteristics and trend analysis of major injuries deaths among children and adolescents in Jiangsu Province from 2012 to 2021].

Objective: To understand the situation and epidemic characteristics of injury deaths among children aged 5 to 24 years in Jiangsu Province from 2012 to 2021 and the trend of annual changes. Methods: The main injury mortality data of children and adolescents was collected, and the crude and standardized mortality rates of road traffic accidents, drowning, suicide, and accidental falls among children and adolescents over a decade and the annual average percentage of change (AAPC) were calculated. The main injury mortality characteristics and trends of children and adolescents of different age groups and genders were analyzed. Results: The total number of injury deaths among 5 to 24 adolescents in Jiangsu Province was 16 052, with a standardized mortality rate of 9.58/100 000. There was no significant trend in the overall standardized mortality rate of injuries (AAPC=-3.450%, P=0.055). The standardized mortality rate of road traffic injuries among children and adolescents showed a decreasing trend over the past decade, with statistical significance (AAPC=-9.406%, P<0.001). The standardized suicide mortality rate showed an upward trend over the past decade, with statistical significance (AAPC=9.000%, P=0.001). The overall injury mortality rate showed an upward trend with age. Suicide rates in males and females were on the rise and both have statistical significance (AAPC=9.420% and AAPC=9.607%, both P<0.05). The standardized mortality rates of female traffic accidents, drowning, and male traffic accidents showed a decreasing trend and were statistically significant (AAPC for female traffic accidents=-7.364%, AAPC for female drowning=-5.352%, and AAPC for male traffic accidents=-10.242%, all P<0.05). The standardized mortality rate of urban and rural traffic accidents showed a decreasing trend and was statistically significant(AAPC=-7.899% and AAPC=-9.421%, both P<0.001). The standardized suicide mortality rate showed an upward trend and statistical significance (AAPC=11.009% and AAPC=7.528%, both P<0.05). Conclusions: The overall injury situation of children and adolescents in Jiangsu Province improved in the past decade from 2012 to 2021, but the suicide mortality rate was on the rise. It is necessary to focus on the mental health issues of this age group and to strengthen the prevention and control of suicide among children and adolescents, in Jiangsu.

[A single-center study on the distribution and antibiotic resistance of pathogens causing bloodstream infection in patients with hematological malignancies].

Objective: To study the incidence of bloodstream infections, pathogen distribution, and antibiotic resistance profile in patients with hematological malignancies. Methods: From January 2018 to December 2021, we retrospectively analyzed the clinical characteristics, pathogen distribution, and antibiotic resistance profiles of patients with malignant hematological diseases and bloodstream infections in the Department of Hematology, Nanfang Hospital, Southern Medical University. Results: A total of 582 incidences of bloodstream infections occurred in 22,717 inpatients. From 2018 to 2021, the incidence rates of bloodstream infections were 2.79%, 2.99%, 2.79%, and 2.02%, respectively. Five hundred ninety-nine types of bacteria were recovered from blood cultures, with 487 (81.3%) gram-negative bacteria, such as Klebsiella pneumonia, Escherichia coli, and Pseudomonas aeruginosa. Eighty-one (13.5%) were gram-positive bacteria, primarily Staphylococcus aureus, Staphylococcus epidermidis, and Enterococcus faecium, whereas the remaining 31 (5.2%) were fungi. Enterobacteriaceae resistance to carbapenems, piperacillin/tazobactam, cefoperazone sodium/sulbactam, and tigecycline were 11.0%, 15.3%, 15.4%, and 3.3%, with a descending trend year on year. Non-fermenters tolerated piperacillin/tazobactam, cefoperazone sodium/sulbactam, and quinolones at 29.6%, 13.3%, and 21.7%, respectively. However, only two gram-positive bacteria isolates were shown to be resistant to glycopeptide antibiotics. Conclusions: Bloodstream pathogens in hematological malignancies were broadly dispersed, most of which were gram-negative bacteria. Antibiotic resistance rates vary greatly between species. Our research serves as a valuable resource for the selection of empirical antibiotics.

Exogenous sodium hydrosulfide can attenuate naloxone-precipitated withdrawal syndromes and affect cAMP signaling pathway in heroin-dependent rat's nucleus accumbens.

BACKGROUND: H2S is a novel type of endogenous neural regulatory factor and gaseous mediator. Exogenous H2S can increase heroin-induced learning and memory damage in rat and alleviates heroin-induced rat hippocampus damage through antioxidant and anti-apoptosis effects. OBJECTIVE: Aim of this study was to identify whether hydrogen sulfide (H2S) protects heroin withdrawal rat is related with adenylate cyclase (AC)-cAMP-protein kinase A (PKA)-cAMP response element-binding protein (CREB) signaling pathway in heroin-dependent rat's nucleus accumbens or not. METHODS: Male Spragne-Dawley rats were randomly divided into Saline + Saline group, Saline + sodium hydrosulfide (NaHS) group, Saline + Heroin group, NaHS + Heroin group according to the principle of increasing heroin dosage day by day, with the establishment of heroin-naloxone-induced withdrawal symptoms determined at day 10. Then the levels of H2S and cAMP and AC and PKA activities were assayed, and the level of phosphorylated CREB (p-CREB), the levels of phosphorylated N-methyl-D-aspartate receptor 1 subunit (p-NR1), phosphorylated N-methyl-D-aspartate receptor 2a subunit (p-NR2A) and phosphorylated N-methyl-D-aspartate receptor 2b subunit (p-NR2B) were assayed in nucleus accumbens. RESULTS: Exogenous H2S can alleviate heroin withdrawal symptoms by increasing the level of H2S level in nucleus accumbens. Exogenous H2S can decrease the high activities of AC, PKA and the high levels of cAMP, p-CREB caused by heroin. Furthermore, exogenous H2S can decrease the high level of p-NR1 and can increase the low levels of p-NR2A and p-NR2B caused by heroin. It is surprising that exogenous H2S treatment alone was able to raise the activities of AC and PKA as well as the levels of cAMP, p-CREB, p-NR1, p-NR2A and p-NR2B. CONCLUSIONS: Exogenous H2S decreases naloxone-precipitated withdrawal signs, maybe through decreasing AC/cAMP/PKA/CREB/NMDR signaling pathway in heroin-dependent rats' nucleus accumbens.

[Measuring laryngotracheal stenosis by extracting centerline based on CT 3D reconstruction].

Objective: To compare the accuracy of the centerline extracted based on CT 3D reconstruction and conventional CT 3D reconstruction in measuring the length and degree of laryngotracheal stenosis. Methods: A retrospective analysis was performed on 35 patients with laryngotracheal stenosis (including 19 cases without tracheotomy and 16 cases with tracheotomy) treated in the Department of Otorhinolaryngology Head and Neck Surgery of the First Affiliated Hospital of Guangxi Medical University from March 2006 to March 2016, including 20 males and 15 females, whose ages ranged from 1 to 73 years, with a median age of 40.5 years. And CT data of 20 normal subjects were included in the same period, including 10 males and 10 females, whose ages ranged from 20 to 63 years, with a median age of 37.0 years. The continuous cross-sectional area of the airway perpendicular to the centerline was obtained by Mimics software. The area was compared with the discontinuous cross-sectional areas reconstructed by conventional CT 3D reconstruction software advantage workstation, also the length of cervical trachea, the length of stenosis, and the minimum airway area were compared. Multi-factor linear stepwise regression method was used to analyze the factors influencing the measuring difference between the two methods. Three patients with laryngotracheal stenosis were selected, and the measured stenosis length was compared with the surgical specimens to evaluate the accuracy of the two methods. SPSS 26.0 software was used for statistical analysis. Results: In normal people, the areas of thyroid cartilage notch, glottis, inferio thyroid cartilage margin, inferio cricoid cartilage margin, and suprasternal notch planes measured by Mimics centerline method were smaller than those measured by conventional CT 3D reconstruction (t thyroid cartilage notch=4.685, tglottis=3.791, tlower thyroid cartilage margin=5.621, tlower cricoid cartilage margin=6.312, tsuprasternal notch plane=6.436, P<0.05). And the airway length measured by Mimics centerline method from the inferior thyroid cartilage to the superior sternal notch was longer (t=9.79, P<0.001). In laryngotracheal stenosis, in the non-tracheotomy group, the minimum airway area measured by Mimics centerline method was smaller and the stenosis length was longer than those measured by the conventional CT 3D reconstruction, and the difference was statistically significant (tminimum airway area=2.562, tstenosis length=5.240, P<0.05). In the tracheotomy group, the stenosis length measured by Mimics centerline method was longer than that measured by conventional CT 3D reconstruction, and the difference was statistically significant (tstenosis length=2.854, P<0.05). Multi-factor linear regression analysis showed that different CT thickness had a statistically significant effect on the difference in the length of stenosis measured by the two methods (b=-5.370, t=-3.306, P=0.004), and different tracheal forward angle had a statistically significant effect on the difference in the minimum airway area measured by the two methods (b=-0.419, t=-2.208, P=0.04). The difference between the measured length of the Mimics centerline method and the intraoperative specimens was less than 0.5 mm. Conclusion: The centerline extracted based on CT 3D reconstruction can precisely reflect the laryngotracheal morphology and measure laryngotracheal stenosis more accurately.

Osteocalcin prevents insulin resistance, hepatic inflammation, and activates autophagy associated with high-fat diet-induced fatty liver hemorrhagic syndrome in aged laying hens.

The aim of this study was to investigate the effects of osteocalcin (OCN) on fatty liver hemorrhagic syndrome (FLHS) in aged laying hens. Thirty 68-week-old White Plymouth laying hens were randomly assigned into conventional single-bird cages, and the cages were randomly allocated into one of 3 treatments (n = 10): normal diet (ND + vehicle, ND + V), high-fat diet (HFD + vehicle, HFD + V), and HFD + OCN (3 µg/bird, 1 time/2 d, i.m.) for 40 d. At day 30, oral glucose tolerance tests (OGTT) and insulin tolerance tests (ITT) were performed. At the end of experiment, the hens were euthanized followed by blood collection. The plasma aspartate transaminase (AST), alkaline phosphatase (ALP), total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) were measured using an automatic biochemistry analyzer. Pathological changes in the liver were examined under both light and transmission electron microscopes. The plasma inflammatory factors including interleukin-1 (IL-1), IL-6, and tumor necrosis factor-alpha (TNF-α) were analyzed by ELISA, and the gene expressions of these inflammatory factors in the liver were analyzed by real-time PCR. The level of oxidative stress was evaluated using malondialdehyde (MDA) and glutathione peroxidase (GSH-Px) assay kits, respectively. The results showed that HFD + V hens had more severe liver hemorrhage and fibrosis than ND + V hens (P < 0.05). The ultramicrostructural examination showed that hepatocytes of HFD + V hens exhibited necrotic pyknosis showing great intracellular electron, mitochondrial swelling, shrunk nucleus, and absence of autolysosomes. Osteocalcin mitigated HFD + V-induced pathological changes in aged laying hens. High-fat diet + OCN hens had higher insulin sensitivity; lower liver concentrations of MDA (P = 0.12) but higher GSH-Px (P < 0.05); and lower blood TNF-α concentrations (P < 0.05) and mRNA expressions (P < 0.05) than HFD + V hens. These results suggest OCN functions in preventing the FLHS process in old laying hens through inhibiting excessive energy diet-induced metabolic disorder, oxidative stress, and related pathological damage.

A Luciferase-EGFP Reporter System for the Evaluation of DNA Methylation in Mammalian Cells.

DNA methylation is an essential epigenetic modification involved in numerous biological processes. Here, we present a cell-based system pLTR-Luc2P-EGFP for evaluation of DNA methylation in mammalian cells. In this system, the expression of reporter gene luciferase2P (Luc2P)-EGFP is under the control of HIV-1 promoter 5' long terminal repeat (LTR), which contains multiple CpG sites. Once these sites are methylated, the expression of Luc2P-EGFP is turned off, which may be visualized under fluorescence microscopy, with quantification performed in luciferase activity assay. As a proof of principle, pLTR-Luc2P-EGFP was methylated in vitro, and transfected into 293T cells, where the reduction of Luc2P-EGFP expression was confirmed. Premixed reporter DNA samples with the methylation levels varying from 0 to 100% were used for quantitative measurements of DNA methylation. The resulting standard curves indicated the accuracy of luciferase activity exceeding that of the Western blotting against EGFP. The Bland-Altman analysis showed that data from luciferase activity assay were in good agreement with the actual DNA methylation levels. In summary, we have established a reporter system coupled with reliable detection technique capable of efficient quantifying the changes in methylation in mammalian cells. This system may be utilized as a high throughput screening tool for identifying molecules that modulate DNA methylation.