Blueberry anthocyanins extract attenuates oxidative stress and angiogenesis on an in vitro high glucose-induced retinopathy model through the miR-33/GLCCI1 axis

Background:Diabetes retinopathy(DR)is a complication of diabetes that affects patients’vision.Previous studies have found blueberry anthocyanins extract(BAE)can inhibit the progression of DR,but its mechanism is not completely clear.Methods:To study the role of BAE in diabetes retinopathy,we treated human retinal endothelial cells(HRCECs)with 30 mM high glucose to simulate the microenvironment of diabetes retinopathy and used BAE to intervene the in vitro high glucose-induced retinopathy model.HRCEC cell viability and apoptosis rates were examined by Cell Counting Kit 8(CCK-8)assay and flow cytometry assay.The binding sites between miR-33 and glucocorticoid-induced transcript 1(GLCCI1)were assessed by luciferase reporter assay.Retinal neovascularization and oxidative stress contribute to diabetic retinopathy.The tubule formation assay was applied to detect the retinal neovascularization.The oxidative stress in the HRCECs was manifested by the reactive oxygen species(ROS)level,the malondialdehyde(MDA)level,and the superoxide dismutase(SOD)activity.Results:Compared with HRCECs cells cultured under normal conditions,high glucose(HG)can induce oxidative stress in HRCRCs,specifically manifested in the increase of ROS and MDA levels,and the decrease of SOD activity.BAE relieved the tubule formation in n the HRCEC.BAE also relieved the ROS and MDA levels and increased the SOD activity.Luciferase reporter assay revealed that GLCCI1 is a target molecule downstream of miR-33.In HRCEC,BAE significantly inhibited the expression of miR-33 induced by HG.miR-33 mimic inhibited the BAE’s effects on oxidative stress and angiogenesis in an in vitro high glucose-induced retinopathy model.Conclusion:BAE alleviated the oxidative stress and microangiogenesis of HRCEC by regulating the miR-33/GLCCI1 axis.

Characterization of blueberry exosome-like nanoparticles and miRNAs with potential cross-kingdom human gene targets

Edible plant derived exosome-like nanoparticles(ELNs)have been shown to have multiple nutraceutical functions.However,the diversity of plant materials makes the plant derived ELN study challenging.More efforts are still needed to explore the feasible isolation methods of edible plant derived ELNs and the possible roles of food-derived ELNs in improving human health.In this study,a size exclusion chromatography based method was compared with the traditional ultracentrifugation method to isolate blueberry derived ELNs(B-ELNs),and the miRNA profile of B-ELNs was analyzed by high-throughput sequencing.A total of 36 miRNAs were found to be enriched in B-ELNs compared with berry tissue,and their potential cross-kingdom human gene targets were further predicted.Results showed that size exclusion chromatography was effective for B-ELN isolation.The most abundant miRNAs in B-ELNs mainly belonged to the miR166 family and miR396 family.Target gene prediction indicated that B-ELNs could potentially regulate pathways related to the human digestive system,immune system and infectious diseases.

Major Diseases and Pests of Blueberry in Guizhou Province and Progress in Their Prevention and Control

As the blueberry industry continues to evolve,the effective control of its diseases and pests has become an essential component of local agricultural development.This paper provides a comprehensive overview of the principal types of blueberry diseases and pests in Guizhou Province,along with the corresponding control measures,in order to serve as a valuable reference for blueberry growers.

Adaptability of Some Blueberry Species in Tianjin

[Objective] The aim was to research the adaptability of 8 blueberry species in Tianjin. [Method] With 8 blueberry species as materials, blueberry was explored in terms of plant characters, the type of fruit-bearing branch, phenological phase, fruit economic characters, and yielding capacity. [Result] Al blueberry species were suitable to be grown in Tianjin with improved soils, showing blueberry growth characters, without any virus or diseases. General y speaking, species B, C, D, E, F and G can be grown in soils exposed to the air as fresh food; species A and H are suitable to be cultivation; species C tastes sour, which can be planted for further production. [Conclusion] The research provides references for blueberry introduction, selection, and cultivation in the region with similar ecological characters.

Study on Grafting and Cultivation of ‘Spartan'Blueberry

The results of grafting improvement and cultivation trials in the Yangtze River Delta of China showed that the rootstock cultivar had a great influence on plant growth and development, fruit yield and quality of ＂Spartan＂ blueberry. Among the seven rootstock-scion combinations, the northern highbush blueberry （Vaccinium corymbosum） cultivar ＂Croaton＇-＇Spartan＇ combination showed well-healed graft unions, large branch amount, early fruit bearing, high yield, excellent fruit quality and almost no tillers. As a kind of rootstock, ＂Croaton＇ was better than non-grafted ＇Spartan＇ and the two rabbiteye blueberry cultivars ＇Premier＇ and ＇Gardenblue＇. The grafting pattern showed no significant stimulating effect on the plant growth and development, fruit yield and quality of ＇Spartan＇ blueberry in this area, but the overall performance of greenwood cleft grafting was better than that of inlaid bud grafting. Suitable rootstock cultivar and grafting pattern significantly improve plant arowth and development, fruit vield and aualitv of ＇Spartan＇ blueberry.

Effect of blueberry on hepatic and immunological functions in mice

BACKGROUND: Conventional drugs used in the treatment and prevention of liver diseases often have side effects, therefore research into natural substances are of significance. This study examined the effects of blueberry on liver protection and cellular immune functions. METHODS: To determine the effects of blueberry on liver protective function, male mice were orally administered blueberry (0.6 g/10 g) or normal saline for 21 days. Hepatic RNA was extracted by Trizol reagent, and the expression of Nrf2, HO-1, and Nqol was determined by real-time RTPCR. Superoxide dismutase (SOD) and malondialdehyde (MDA) in liver homogenate were determined, and liver index was measured. To assess the effects of blueberry on cellular immune function, male mice received blueberry (0.4, 0.6, or 0.8 g/10 g) for 35 days, and the percentages of CD3(+), CD41(+), and CDS(+) T lymphocyte subgroups in peripheral blood were detected by flow cytometry, the index of the thymus and spleen was measured, and lymphocyte proliferation in the spleen was determined by MTT assay. RESULTS: Blueberry treatment significantly increased the expression of Nrf2, HO-1, and Nqol, the important antioxidant components in the liver. Hepatic SOD in the blueberry group was higher and MDA was lower than that in the control group (P<0.05). Blueberry also increased the index of the spleen and enhanced the proliferation of lymphocytes of the spleen (P<0.05). The percentages of the CD3(+) and CDA(+) T lymphocyte subsets and the CD4(+)/CD8(+) ratio were also increased by blueberry (P<0.05). CONCLUSIONS: Blueberry induces expression of Nrf2, HO-1, and Nqol, which can protect hepatocytes from oxidative stress. In addition, blueberry can modulate T-cell function in mice. (Hepatobiliary Pancreat Dis Int 2010; 9: 164-168)

Effects of blueberry on hepatic fibrosis and transcription factor Nrf2 in rats

AIM:To investigate the effects of blueberry on hepatic fibrosis and NF-E2-related factor 2(Nrf2) transcription factor in rats.METHODS:Forty-five male Sprague-Dawley rats were randomly divided into control group(A);CCl4-induced hepatic fibrosis group(B);blueberry prevention group(C);Dan-shao-hua-xian capsule(DSHX) prevention group(D);and blueberry + DSHX prevention group(E).Liver fibrosis was induced in rats by subcutaneous injection of CCl4 and a high-lipid/low-protein diet for 8 wk(except the control group).The level of hyaluronic acid(HA) and alanine aminotransferase(ALT) in serum was examined.The activity of superoxide dismutase(SOD),glutathione-S-transferase(GST) and malondialdehyde(MDA) in liver homogenates was determined.The degree of hepatic fibrosis was evaluated by hematoxylin and eosin and Masson staining.Expression of Nrf2 and NADPH quinone oxidoreductase 1(Nqo1) was detected by real-time reversed transcribed-polymerase chain reaction,immunohistochemical techniques,and western blotting.RESULTS:Compared with group B,liver indices,levels of serum HA and ALT of groups C,D and E were reduced(liver indices:0.038 ± 0.008,0.036 ± 0.007,0.036 ± 0.005 vs 0.054 ± 0.009,P<0.05;HA:502.33 ± 110.57 ng/mL,524.25 ± 255.42 ng/mL,499.25 ± 198.10 ng/mL vs 828.50 ± 237.83 ng/mL,P<0.05;ALT:149.44 ± 16.51 U/L,136.88 ± 10.07 U/L,127.38 ± 11.03 U/L vs 203.25 ± 31.62 U/L,P<0.05),and SOD level was significantly higher,but MDA level was lower,in liver homogenates(SOD:1.36 ± 0.09 U/mg,1.42 ± 0.13 U/mg,1.50 ± 0.15 U/mg vs 1.08 ± 0.19 U/mg,P<0.05;MDA:0.294 ± 0.026 nmol/mg,0.285 ± 0.025 nmol/mg,0.284 ± 0.028 nmol/mg vs 0.335 ± 0.056 nmol/mg,P<0.05).Meanwhile,the stage of hepatic fibrosis was significantly weakened(P<0.05).Compared with group A,the activity of GST liver homogenates and expression levels of Nrf2 and Nqo1 in group B were elevated(P<0.05).The expression level of Nrf2 and Nqo1 in groups C,D,and E were increased as compared with group B,but the difference was not significant.CONCLUSION:Blueberry has preventive and protective effects on CCl4-induced hepatic fibrosis by reducing hepatocyte injury and lipid peroxidation.However,these effects may not be related to the activation of Nrf2 during long-term of CCl4.

High-throughput sequencing of highbush blueberry transcriptome and analysis of basic helix-loop-helix transcription factors

The highbush blueberry（Vaccinium corymbosum）,Duke,was used to construct a de novo transcriptome sequence library and to perform data statistical analysis.Mega 4,CLC Sequence Viewer 6 software,and quantitative PCR were employed for bioinformatics and expression analyses of the basic helix-loop-helix（BHLH）transcription factors of the sequencing library.The results showed that 28.38 gigabytes of valid data were obtained from transcriptome sequencing and were assembled into 108 033 unigenes.Functional annotation showed that 32 244 unigenes were annotated into Clusters of Orthologous Groups（COG）and Gene Ontology（GO）databases,whereas the rest of the 75 789 unigenes had no matching information.By using COG and GO classification tools,sequences with annotation information were divided into 25 and 52 categories,respectively,which involved transport and metabolism,transcriptional regulation,and signal transduction.Analysis of the transcriptome library identified a total of 59 BHLH genes.Sequence analysis revealed that 55 genes of that contained a complete BHLH domain.Furthermore,phylogenetic analysis showed that BHLH genes of blueberry（Duke）could be divided into 13 sub-groups.PCR results showed that 45 genes were expressed at various developmental stages of buds,stems,leaves,flowers,and fruits,suggesting that the function of BHLH was associated with the development of different tissues and organs of blueberry,Duke.The present study would provided a foundation for further investigations on the classification and functions of the blueberry BHLH family.

Cyanidin-3-O-galactoside and Blueberry Extracts Supplementation Improves Spatial Memory and Regulates Hippocampal ERK Expression in Senescence-accelerated Mice

Objective To investigate whether the antioxidation and the regulation on the Extracellular Regulated Protein Kinases （ERK） signaling pathway are involved in the protective effects of blueberry on central nervous system. Methods 30 Senescence-accelerated mice prone 8 （SAMP8） mice were divided into three groups and treated with normal diet, blueberry extracts （200 mg/kg.bw/day） and cyaniding-3-O-galactoside （Cy-3-GAL） （50 mg/kg.bw/day） from blueberry for 8 weeks. 10 SAMR1 mice were set as control group. The capacity of spatial memory was assessed by Passive avoidance task and Morris water maze. Histological analyses on hippocampus were completed. Malondialdehyde （MDA） levels, Superoxide Dismutase （SOD） activity and the expression of ERK were detected. Results Both Cy-3-GAL and blueberry extracts were shown effective functions to relieve cellular injury, improve hippocampal neurons survival and inhibit the pyramidal cell layer damage. Cy-3-GAL and blueberry extracts also increased SOD activity and reduced MDA content in brain tissues and plasma, and increased hippocampal phosphorylated ERK （p-ERK） expression in SAMP8 mice. Further more, the passive avoidance task test showed that both the latency time and the number of errors were improved by Cy-3-GAL treatment, and the Morris Water Maze test showed significant decreases of latency were detected by Cy-3-GAL and blueberry extracts treatment on day 4. Conclusion Blueberry extracts may reverse the declines of cognitive and behavioral function in the ageing process through several pathways, including enhancing the capacity of antioxidation, altering stress signaling. Cy-3-GAL may be an important active ingredient for these biological effects.

Comparison of Different Cultivars of Blueberry Overwintering Ability in Qingdao of China

In order to select severe overwintering abilitied blueberry cultivars for cold resistance of annual branches, promoting breeding high quality cultivars, a study had been conducted into field-planting shoots overwintering ability and the relative conductivity, MDA content, SOD of 7 different kinds of blueberries under artificial cooling process. According to survey results, under field conditions, Different cultivars of blueberries showed significantly difference, ranging from 56.67% of Bluegold to 12.80% of Darrow. With the temperature decreasing, changes of relative conductivity, MDA content and SOD in annual branches had a strong regularity and the hardiness of these cultivars is: Northland > Chippewa > Coville > Bluecroup > Darrow > Bluegold > Powderblue.

Validation of a SPME-GC Method for Azoxystrobin and Pyraclostrobin in Blueberries, and Their Degradation Kinetics

Analytical method for the residues of strobilurins azoxystrobin and pyraclostrobin in blueberries was developed. Fungicide residues were determined by solid-phase microextraction (SPME) cou-pled to gas chromatography with micro-electron capture detector. The effect of pH values and fiber coatings were studied. The SPME fiber coating selected was 100 μm PDMS. The method is selective with adequate precision and high accuracy and sensitivity. Recoveries ranged within the 100% - 106% range for azoxystrobin, and 96% - 106% range for pyraclostrobin;and detection and quantification limits were 2.0 and 6.0 μg/kg for azoxystrobin, and 26.0 and 86.0 μg/kg for py-raclostrobin, respectively. Statistical parameters indicated the occurrence of matrix effect;con-sequently calibration was performed on spiked samples. Degradation of azoxystrobin and pyra-clostrobin was studied in blueberry fields located in Concordia, Argentina, with fruits from Emerald and Jewel varieties. The degradation of these fungicides in both blueberry varieties studied followed a first order rate kinetics, and the half-life for azoxystrobin was 11.6 and 17.8 days for Emerald and Jewel cultivars;and for pyraclostrobin was 5.5 and 8.0 days, respectively.

Evaluation of sugar and organic acid composition and their levels in highbush blueberries from two regions of China

Sweet and sour are the most important taste of blueberries,and they are produced by sugar and acid,respectively.Their contributions to the taste depend not only on the levels of sugar and acid,but also on the types and relative proportions of sugar and acid.Therefore,it is very important to evaluate the composition and levels of sugar and acid in blueberries.Regional differences and variety diversity also affect the sugar and acid characteristics of fruits.Therefore,this study selected two main producing regions in northern China(Weihai and Yingkou)to examine the sugar and acid characteristics of 11 common blueberry cultivars.The indexes measured included soluble sugars,organic acids,soluble solid content and titratable acidity.The results showed that glucose and fructose were the major sugars,and citric acid and quinic acid were the major organic acids.Correlation analysis showed that glucose,fructose,and sucrose were positively correlated with total sugar content;the citric acid content was positively correlated with the titratable acidity and total organic acids.Titratable acidity,glucose,fructose,sucrose,total sugar content,citric acid,shikimic acid and total acid content of the blueberries varied significantly between regions(P<0.05).In general,compared with Weihai blueberries,Yingkou blueberries had higher sugar content and lower acid content.The results of this study may provide useful references for the evaluation of sweet and sour flavors and cultivar selection of blueberries.

Effect of Soil p H on the Growth,Photosynthesis and Mineral Element Content in Own-Rooted Saplings and Grafted Saplings of Blueberry

The adaptation of blueberry to higher soil p H is poor. To explore the effect and probable mechanism of the adaptation of grafted blueberry( Vaccinium spp.) saplings to the soil environment with higher p H value. We took three types of saplings,( i) own-rooted Wufanshu( V. bracteatum),an economical shrub belonging to the same genus and family with blueberry,also known as oriental blueberry,( ii) own-rooted‘Sharpblue',a southern highbush blueberry( V. corymbosum) cultivar,and( iii) grafted-bluebedrry as experimental materials,which were grown in quartz sand medium for 90 d,irrigated with nutrient solution of three levels of p H values,respectively,i. e. p H 4. 6,p H 5. 8 and p H7. 0. Results showed that root development,shoot growth and dry biomass accumulation of grafted blueberry were significantly better than those of own-rooted blueberry grown in higher p H medium; compared with own-rooted blueberry saplings,leaf chlorophyll content,chlorophyll a/b ratio,chlorophyll fluorescence chemical efficiency and photosynthetic efficiency of grafted blueberry saplings grown in higher p H medium were all significantly improved. Results also showed that mineral element content of leaves,especially Mg and Fe elemental content of grafted blueberry was significantly higher than that of own-rooted blueberry saplings grown in higher p H media. Grafting-propagation of blueberry saplings taking Wufanshu as rootstock significantly enhanced the adaptation of blueberry to soil environment with higher p H value.

Current Development Situation and Prospects of Blueberry in Guangdong Province

With high nutrition and health value,blueberry receives much concern in recent years. Blueberry experts in the world are cultivating new blueberry varieties. They have come up with constructive theoretical support and planting technology researches and are constantly devoted to expanding blueberry planting latitudes. This paper elaborated researches of blueberry at home and abroad,current development situation of blueberry in China and Guangdong Province,analyzed strengths of planting blueberry in Guangdong. Besides,it came up with recommendations for development of blueberry industry. Finally,it discussed development prospects of blueberry planting in Guangdong.

Species-specific PCR-based assays for identification and detection of Botryosphaeriaceae species causing stem blight on blueberry in China

Botryosphaeriaceae species are important causal agents of blueberry stem blight worldwide. Blueberry stem blight has become an important disease, potentially affecting the quality and production of blueberries in China. It is difficult and time-consuming to identify at the species level using morphological methods. The aim of this study was to develop polymerase chain reaction（PCR） assays for the diagnosis and early detection of latent infections of blueberry stems by Botryosphaeria spp. Species-specific primers, based on the ribosomal DNA internal transcribed spacer region and β-tubulin gene, were designed and selected for use in PCR assays. Three primer pairs, Lt347-F/R for Lasiodiplodia theobromae, Np304-F/R for Neofusicoccum parvum and FaF/Bt2b for Botryosphaeria dothidea, successfully amplified specific PCR fragments of different sizes on pure cultures or from blueberry stems inoculated and naturally infected blueberry plants with three pathogens, respectively. These primers did not amplify any PCR fragments from other blueberry stem disease-associated pathogens, such as Phomopsis spp. and Pestalotiopsis spp. This PCR protocol could detect as low as 1 00 pg to 1 ng of purified fungal DNA. This PCR-based protocol could be used for the diagnosis and detection of these pathogens from pure cultures or from infected blueberry plants.

Research on Production Conditions of Highgrade Wine from Northern Blueberry

In order to develop the best brewing condition for high-grade fruit wine from northern blueberry, on the basis of reviewing literatures about blueberry wine manufacturing process, this study put forward a new technological process, including raw materials-juicing-addition of auxiliary materials including pectinase-primary fer- mentation with yeast-post-fermentation-hot and cold treatment-sterilization-packaging- finished product, and sensory indexes, physicochemical indexes and hygienic index- es of the product were inspected according to corresponding national standards and industry standards. The results showed that for northern blueberry pulp, the optimal addition amount of yeast was 1.1 g/L, the fermentation temperature was 22 ℃, and the addition amounts of pectinase and sulfurous acid were 0.3 ml/kg and 100 ppm, respectively; the alcohol degree of the finished product was adjusted to 15.6°; and alternated cold and heat treatment used instead of conventional clarifying agent for removing colloid-like impurities resulted in the brewed product with good wine fra- grance, taste and color.

An Approach for Micropropagation of Blueberry (<i>Vaccinium corymbosum</i>L.) Plants Mediated by Temporary Immersion Bioreactors (TIBs)

A new procedure for blueberry (Vaccinium corymbosum L.) micropropagation in programmed Temporary Immersion Bioreactors (TIBs based on two separate bottles) was developed for the commercial genotypes Biloxi, Sharp Blue and Brillita. Plant cultures were developed in a controlled environment with 0.4 MPa CO2 enrichment, sucrose-reduced medium, and light intensity of 60 mM m-2·s-1. Principal component analysis showed that component 1 (C1) grouped 64.08% of the total variability, while the first two components accounted for 86.97%. Representation of the principal components demonstrated three clusters corresponding with the blueberry genotypes, and within each cluster plants micropropagated in agar-base medium grouped separately from those plants multiplied in TIBs. Both plant number and total internodes traits (related to the productive efficiency) were demonstrated superior in blueberries propagated in TIBs. Additionally, when transferred to greenhouse conditions, blueberries propagated in TIBs showed higher adaptability and growing rates than those cultured by the conventional approach, altogether evidencing the occurrence of a photomixotrophic stage in the vitroplantlets cultured in TIBs.

Initial Screening of 10 Plant Essential Oils in Vitro Antibacterial Activity Against Blueberry Pathogens

In order to develop natural antistaling agent for blueberries preservation,the antibacterial activities of 10 kinds of plant essential oils were analyzed by oxford-cup test and tube dilution method,and their inhibitory abilities to two kinds of representative Gram bacteria which are Escherichia coli and Staphylococcus aureus,and three kinds of harmful fungi in blueberries,involving Alternaria alternata,Penicillium digitatum and Botrytis cinerea,were tested.The results showed that 7 kinds of essential oils can inhibit the three fungi,among which cinnamon oil,clove oil,anise oil and thyme oil have strong fungi static efficacy.Cinnamon oil and thyme oil can effectively inhibit the Gram-positive bacterium(Staphylococcus aureus);cinnamon oil and masson pine oil are strongly inhibitory to Gram-negative bacterium(E.coli).Cinnamon oil has broad-spectrum ability to inhibit fungi,Gram-negative and positive bacteria,with a minimum inhibitory concentration(MIC)ranging from 0.0391 to 0.1560 L/mL.With strong inhibition effects on fungi,cinnamon oil can be used as a compound base oil and mixed with other essential oils to retain the freshness of blueberries,thus achieving better antibacterial and anti-corrosion effects.

Influence of Blueberry and Jaboticaba Agroindustrial Residue Particle Size on Color Change of Corn Starch Based Films Submitted to Different pH Values Solutions

Corn starch,glycerol and agroindustrial residues were used to produce films by casting.By-products from juice processing,blueberry and jaboticaba in powder with different particle sizes were added in the filmogenic matrix to evaluate its potential as a colorimetric indicator.Blueberry and jaboticaba peels are commonly discarded although contain high amount of important compounds as anthocyanins.These compounds have the ability to color change after immersion in different pH values,demonstrating its potential for the intelligent packaging development.Analyses were performed in a colorimeter after films immersion in different buffer solutions.Visual color changes were perceived;reddish and bluish color in acidic and basic pH values,respectively.Independently of the particle size,theΔE* values were greater than 5,showing visually perceptible change to the human eye.The results indicate the potential of use of these residues as a pH indicator for the development of renewable and biodegradable sensor of food deterioration.

Effect of GA_(3) Concentration and Soaking Time on Seed Germination Characteristics of Blueberry

This study aims to explore the effect of GA_(3) concentration and soaking time on seed germination characteristics of blueberry.To be specific,big(B),medium-sized(M),and small(S)blueberry seeds were soaked in 0,100,200,and 300 mg/L GA_(3) solution for 18 h and 200 mg/L GA_(3) solution for 0,24,and 48 h,respectively.The results showed that 300 mg/L GA_(3) had significant effect on the germination of B and M seeds(P<0.01)and enabled the germination 4~5 d and 7~8 d earlier,respectively.Soaking with 200 mg/L GA_(3) for 24 h was conducive to the germination of B and M seeds(P<0.05).However,GA_(3) concentration and soaking time had no obviously promoting effect on the germination of S seeds.The interactions between GA_(3) concentration and seed size and between soaking time and seed size were significant(P<0.01).