Endogenous biosynthesis of docosahexaenoic acid(DHA)regulates fish oocyte maturation by promoting pregnenolone production

Omega-3 polyunsaturated fatty acids(n-3 PUFAs),particularly docosahexaenoic acid(22:6n-3,DHA),play crucial roles in the reproductive health of vertebrates,including humans.Nevertheless,the underlying mechanism related to this phenomenon remains largely unknown.In this study,we employed two zebrafish genetic models,i.e.,elovl2^(-/-)mutant as an endogenous DHAdeficient model and fat1(omega-3 desaturase encoding gene)transgenic zebrafish as an endogenous DHA-rich model,to investigate the effects of DHA on oocyte maturation and quality.Results show that the elovl2^(-/-)mutants had much lower fecundity and poorer oocyte quality than the wild-type controls,while the fat1 zebrafish had higher fecundity and better oocyte quality than wildtype controls.DHA deficiency in elovl2^(-/-)embryos led to defects in egg activation,poor microtubule stability,and reduced pregnenolone levels.Further study revealed that DHA promoted pregnenolone synthesis by enhancing transcription of cyp11a1,which encodes the cholesterol side-chain cleavage enzyme,thereby stabilizing microtubule assembly during oogenesis.In turn,the hypothalamic-pituitary-gonadal axis was enhanced by DHA.In conclusion,using two unique genetic models,our findings demonstrate that endogenously synthesized DHA promotes oocyte maturation and quality by promoting pregnenolone production via transcriptional regulation of cyp11a1.

Impact of stage-specific limb function exercises guided by a selfmanagement education model on arteriovenous fistula maturation status

BACKGROUND The exercise of limb function is the most economical and safe method to promote the maturation of arteriovenous fistula(AVF).However,due to the lack of a uni-fied exercise standard in China,many patients have insufficient awareness of the importance of AVF,leading to poor effectiveness of limb function exercise.The self-management education model can effectively promote patients to take pro-active health-related actions.This study focuses on the characteristics of patients during the peri-AVF period and conducts a phased limb function exercise under the guidance of the self-management education model to observe changes in fac-tors such as the maturity of AVF.AIM To assess the impact of stage-specific limb function exercises,directed by a self-management education model,on the maturation status of AVFs.METHODS This study is a randomized controlled trial involving 74 patients with forearm AVFs from the Nephrology Department of a tertiary hospital in Sichuan Province,China.Patients were randomly divided into an observation group and a control group using a random number table method.The observation group underwent tailored stage-specific limb func-tion exercises,informed by a self-management education model which took into account the unique features of AVF at various stages,in conjunction with routine care.Conversely,the control group was given standard limb function exercises along with routine care.The assessment involves the maturity of AVFs post-intervention,post-operative complications,and the self-management level of the fistula in both groups patients.Analyses were conducted using SPSS version 23.0.Count data were represented by frequency and percentage and subjected to chi-square test comparisons.Measurement data adhering to a normal distribution were presented as mean±SD.The independent samples t-test was utilized for inter-group comparisons,while the paired t-test was used for intra-group comparisons.For measurement data not fitting a normal distribution,the median and interquartile range were presented and analyzed using the Wilcoxon rank sum test.RESULTS At the 8-wk postoperative mark,the observation group demonstrated significantly higher scores in AVF symptom recognition,symptom prevention,and self-management compared to the control group(P<0.05).However,the variance in symptom management scores between the observation and control groups lacked statistical signi-ficance(P>0.05).At 4 wk after the operation,the observation group displayed a superior vessel diameter and depth from the skin of the drainage vessels in comparison to the control group(P<0.05).While the observation group did manifest elevated blood flow rates in the drainage vessels relative to the control group,this distinction was not statistically significant(P>0.05).By the 8-wk postoperative interval,the observation group outperformed the control group with notable enhancements in blood flow rates,vessel diameter,and depth from the skin of drainage vessels(P<0.01).Seven days following the procedure,the observation group manifested significantly diminished limb swelling and an overall reduced complication rate in contrast to the control group(P<0.05).The evaluation of infection,thrombosis,embolism,arterial aneurysm stenosis,and incision bleeding showed no notable differences between the two groups(P>0.05).By the 4-wk postoperative juncture,complications between the observation and control groups were statistically indistinguishable(P>0.05).CONCLUSION Stage-specific limb function exercises,under the guidance of a self-management education model,amplify the capacity of AVF patients to discern and prevent symptoms.Additionally,they expedite AVF maturation and miti-gate postoperative limb edema,underscoring their efficacy as a valuable method for the care and upkeep of AVF in hemodialysis patients.

Protective effect of resveratrol against cadmium-induced toxicity on ovine oocyte in vitro maturation and fertilization

Background:Heavy metal cadmium(Cd)is a widespread environmental contaminant with a potential toxicity that might negatively affect female reproduction and fertility.It has been reported that Cd exposure impaired the quality of oocytes and led to a defective maturation and fertilization,through oxidative stress induction.Resveratrol(Res)is a natural polyphenol with strong antioxidant properties that exhibited protective role in preventing oocyte redox homeostasis disruption and quality decline.Here,we explored whether the addition of Res to in vitro maturation(IVM)medium might act as a protection against Cd-induced toxicity on ovine oocyte maturation and fertilization.Firstly,we evaluated the effect of supplementing IVM medium with two different Res concentrations(1and 2μmol/L)on nuclear maturation and fertilization of oocytes matured under CdCl2(2μmol/L)exposure.Therefore,the concentration of 1μmol/L Res was selected to analyse the effects of this compound on intracellular ROS levels,mitochondrial(mt)distribution and activity,chromatin configuration,cytoskeleton morphology,cortical granules(CGs)distribution and mRNA expression of genes associated with cellular response to oxidative stress(i.e.SIRT1,SOD 1,GPX1,GSR,CAT)in Cd-exposed in vitro matured oocytes.Results:We found that 1μmol/L Res restored the reduced oocyte meiotic competence induced by Cd exposure as well as,Res sustained oocyte ability to be normally fertilized and decreased polyspermic fertilization at both tested concentrations.Moreover,we demonstrated that 1μmol/L Res mitigated Cd-induced alterations of oocyte cytoplasmic maturation by reducing reactive oxygen species(ROS)accumulation,preventing mt dysfunction,maintaining the correct meiotic spindle and cortical F-actin assembly and the normal cortical granule distribution as well as up-regulating SIRT1,SOD1 and GPX1 genes.Conclusions:Taken together,our findings highlighted the beneficial influence exerted by Res in preventing Cdinduced disturbance of nuclear and cytoplasmic maturation and subsequent fertilization in ovine oocytes.Res treatment may help to establish defence strategies counteracting Cd-induced toxicity on the female gamete.

Neurovascular unit on a chip:the relevance and maturity as an advanced in vitro model

The brain is a high-energy demanding organ,consuming around 20%of the metabolic energy generated.To fulfill this demand,cerebral blood flow(CBF)supplies oxygen and glucose continuously through the intricate network of cerebral blood vessels.Although for many years brain activity and blood flow were conceived as independent processes,MRI-based functional brain imaging demonstrated that there is a coupling between them.

Enhancer of Shoot Regeneration 2(ESR2)regulates pollen maturation and vitality in watermelon(Citrullus lanatus)

Watermelon(Citrullus lanatus)holds global significance as a fruit with high economic and nutritional value.Exploring the regulatory network of watermelon male reproductive development is crucial for developing male sterile materials and facilitating cross-breeding.Despite its importance,there is a lack of research on the regulation mechanism of male reproductive development in watermelon.In this study,we identified that ClESR2,a VIIIb subclass member in the APETALA2/Ethylene Responsive Factor(AP2/ERF)superfamily,was a key factor in pollen development.RNA insitu hybridization confirmed significantClESR2 expression in the tapetum and pollen during the later stage of anther development.The pollens of transgenic plants showed major defects in morphology and vitality at the late development stage.The RNA-seq and protein interaction assay confirmed that ClESR2 regulates pollen morphology and fertility by interacting with key genes involved in pollen development at both transcriptional and protein levels.These suggest that Enhancer of Shoot Regeneration 2(ESR2)plays an important role in pollen maturation and vitality.This study helps understand the male reproductive development of watermelon,providing a theoretical foundation for developing male sterile materials.

Effect of thermal maturation and organic matter content on oil shale fracturing

The Pabdeh Formation represents organic matter enrichment in some oil fields,which can be considered a source rock.This study is based on the Rock–Eval,Iatroscan,and electron microscopy imaging results before and after heating the samples.We discovered this immature shale that undergoes burial and diagenesis,in which organic matter is converted into hydro-carbons.Primary migration is the process that transports hydrocarbons in the source rock.We investigated this phenomenon by developing a model that simulates hydrocarbon generation and fluid pressure during kerogen-to-hydrocarbon conversion.Microfractures initially formed at the tip/edge of kerogen and were filled with hydrocarbons,but as catagenesis progressed,the pressure caused by the volume increase of kerogen decreased due to hydrocarbon release.The transformation of solid kerogen into low-density bitumen/oil increased the pressure,leading to the development of damage zones in the source rock.The Pabdeh Formation’s small porethroats hindered effective expulsion,causing an increase in pore fluid pressure inside the initial microfractures.The stress accumulated due to hydrocarbon production,reaching the rock’s fracture strength,further contributed to damage zone development.During the expansion process,microfractures preferentially grew in low-strength pathways such as lithology changes,laminae boundaries,and pre-existing microfractures.When the porous pressure created by each kerogen overlapped,individual microfractures interconnected,forming a network of microfractures within the source rock.This research sheds light on the complex interplay between temperature,hydrocarbon generation,and the development of expulsion fractures in the Pabdeh Formation,providing valuable insights for understanding and optimizing hydrocarbon extraction in similar geological settings.

CRISPR/Cas9-mediated knockout of E4 gene promotes maturation in soybean

Soybean is a broadly popular and extensively cultivated crop,however,many high-yield and high-quality varieties require specific growth conditions,restricting their widespread adoption.The appropriate light conditions and photoperiod must be attained for these varieties to thrive in new environments.In this study,we employed CRISPR/Cas9 to design two sgRNAs aimed at knocking out the maturity-related gene E4 in a major American soybean variety called''Jack'',which belongs to maturity group MGII.E4 gene is primarily involved in the photoperiodic flowering and maturity in soybean,making it an ideal candidate for genetic manipulation.We successfully obtained 1 homozygous E4-SG1 mutant type with 1-bp insertion,and 4 homozygous E4-SG2 mutants type with 2-bp deletion,7-bp deletion,61-bp deletion,and 1-bp insertion,respectively.The homozygous e4 mutant plants contained early termination codons devoid of transgenic elements.Additionally,no potential offtarget sites of the E4 gene were detected.A comparative analysis revealed that,unlike the wild-type,the maturity time of homozygous e4 mutants was early under both short-day and long-day conditions.These mutants offer novel germplasm resources that may be used to modify the photoperiod sensitivity and maturity of soybean,enhancing its adaptability to high-latitude regions.

A New Micropropagation Technology of Tilia amurensis:In VitroMicropropagation of Mature Zygotic Embryos and the Establishment of a PlantRegeneration System

Tilia amurensis is an economically valuable broadleaf tree species in Northeast China.The production of highqualityT.amurensis varieties at commercial scales has been greatly limited by the low germination rates.Thereis thus a pressing need to develop an organogenesis protocol for in vitro propagation of T.amurensis to alleviate ashortage of high-quality T.amurensis seedlings.Here,we established a rapid in vitro propagation system forT.amurensis from mature zygotic embryos and analyzed the effects of plant growth regulators and culture mediain different stages.We found that Woody plant medium(WPM)was the optimal primary culture medium formature zygotic embryos.The highest callus induction percentage(68.76%)and number of axillary buds induced(3.2)were obtained in WPM+0.89μmol/L 6-benzyladenine(6-BA)+0.46μmol/L kinetin(KT)+0.25μmol/Lindole-3-butryic acid(IBA)+1.44μmol/L gibberellin A_(3)(GA_(3)).The multiple shoot bud development achievedthe highest percentage(83.32%)in the Murashige and Skoog(MS)+2.22μmol/L 6-BA+0.25μmol/L IBA+1.44μmol/L GA_(3).The rooting percentage(96.70%)was highest in 1/2 MS medium+1.48μmol/L IBA.Thesurvival percentage of transplanting plantlets was 82.22%in soil:vermiculite:perlite(5:3:1).Our study is the firstto establish an effective organogenesis protocol for T.amurensis using mature zygotic embryos.

Effects of in Vitro Maturation Time of Oocytes on Sheep Nuclear Transfer Efficiency

[Objective] The study aimed to provide references for the time of oocyte maturation in vitro and enucleation in the course of sheep nuclear transfer(NT).[Method] Compared the effects of different maturation time of oocytes on enucleation efficiency and reconstructed embryo development by means of blind enucleation and fluorescence microscopy.[Result] Treatment of IVM(in vitro maturation)19-21 h was significantly higher than IVM 16-18 h treatment in oocyte maturation rate(P<0.05)and was significantly higher than IVM 22-24 h treatment in enucleation rate(P<0.05).Three treatments had no significant difference in cleavage rate and blastocyst rate(P>0.05),but IVM 19-21 h treatment was significantly higher than the other 2 treatments in average cell number of blastocysts(P<0.05).[Conclusion] The appropriate in vitro maturation time of oocytes was 19-21 h for sheep nuclear transfer,which could significantly improve the quality of blastocysts according to the cell number per blastocyst(P<0.05).

Improvement of in vitro Cytoplasmic Maturation of Denuded Porcine Oocytes by Coculture with Follicular Mural Granulosa Cells

[Objective] This study aimed to improve the in vitro maturation quality of denuded porcine oocytes and provide scientific basis for establishing a stable and efficient denuded oocyte culture system. [Method] The first polar body extrusion rate, oocyte glutathione （GSH） content, positive rate of brilliant cresyl blue （BCB） staining and development potential of activated oocytes or fertilized oocytes were employed as main indicators to investigate the effects of follicular mural granulosa cell （MGC） coculture on cytoplasmic maturation of cumulus cell-removal oocytes （Denuded Oocyte, DO）. [Result] According to in vitro maturation results, compared with DO group, the first polar body extrusion rate of porcine oocytes in DO＋MGC group was not significantly different, but the nuclear maturation process was improved and was more similar to that in COC （cumulus-oocyte complex） group. Detection of GSH content in mature oocytes showed that there was no significant difference between DO＋ MGC group （optical density of 1 053.67） and COC group （optical density of 1 426.00） or between DO＋MGC group and COC＋GC group （optical density of 1 541.00）, however, GSH content in mature oocytes of DO group （optical density of 724.67） was significantly lower than that of COC group and COC＋GC group （P0.05）. Detection of glucose-6-phosphate dehydrogenase （G6PDH） activity showed that there was no significant difference in BCB positive oocyte rate between DO ＋MGC group （88.26% ） and COC group （92.75%） or between DO＋MGC group and DO group （82.86% ）, however, BCB positive oocyte rate of DO group was significantly lower than that of COC group （P0.05）. Furthermore, the cleavage rate and blastocyst rate of activated mature oocytes derived from DO ＋MGC group （94.98% and 43.67% , respectively） were significantly higher than those from DO group （52.54% and 8.97%, respectively） （P0.05）, and were not significantly different compared with those from COC group （97.11% and 38.30%, respectively）. In addition, the cleavage rate of fertilized oocytes derived from DO＋MGC group （72.65%） showed no significant difference compared with that from DO group （63.59%）, but the blastocyst rate of DO＋MGC group was significantly higher than that of DO group （9.88%） （P0.05）. [Conclusion] MGC coculture can significantly improve the in vitro cytoplasmic maturation quality of denuded porcine oocytes, thereby enhancing the subsequent developmental potential.

In vitro Maturation of Tan Sheep Oocytes

[Objective] This study aimed to investigate the appropriate concentrations of follicle-stimulating hormone（FSH）, luteotropic hormone（LH） and estrodiol（E2） during in vitro maturation of Tan sheep oocytes. [Method] Tan sheep oocytes were divided into five groups for in vitro maturation culture： control group, FSH group（10,50, 100, 200 and 300 μg/ml FSH, respectively）, LH group（5, 10, 20, 50 and 100μg/ml LH, respectively）, E2group（5, 10, 25, 50 and 100 μg/ml E2, respectively）, and FSH ＋ LH group（100 μg/ml FSH ＋ 20 μg/ml LH）. The releasing rate of first polar bodies was analyzed. [Result] The maturation rate of Tan sheep oocytes in 100 μg/ml FSH ＋ 20 μg/ml LH group reached the highest（64.64%）, which was significantly higher than that in other four groups（P〈0.05）; among different FSH concentrations,100 μg/ml FSH was superior to other four concentrations and the control group, exhibiting significant differences（P〈0.05）; among different LH concentrations, 20 μg/ml LH was superior to other four concentrations and the control group, exhibiting significant differences（P〈0.05）; among different E2 concentrations, 50 μg/ml E2 was superior to other four concentrations and the control group, exhibiting significant differences（P〈0.05）. [Conclusion] Under the experimental conditions, 100 μg/ml FSH ＋20 μg/ml LH was the most appropriate hormone combination for in vitro maturation of Tan sheep oocytes.

Selection of oocytes for in vitro maturation by brilliant cresyl blue staining： a study using the mouse model

Selecting oocytes that are most likely to develop is crucial for in vitro fertilization and animal cloning. Brilliant cresyl blue （BCB） staining has been used for oocyte selection in large animals, but its wider utility needs further evaluation. Mouse oocytes were divided into those stained （BCB＋） and those unstained （BCB-） according to their ooplasm BCB coloration. Chromatin configurations, cumulus cell apoptosis, cytoplasmic maturity and developmental competence were compared between the BCB＋ and BCB- oocytes. The effects of oocyte diameter, sexual maturity and gonadotropin stimulation on the competence of BCB＋ oocytes were also analyzed. In the large- and medium-size groups, BCB＋ oocytes were larger and showed more surrounded nucleoli （SN） chromatin configurations and higher frequencies of early atresia, and they also gained better cytoplasmic maturity （determined as the intracellular GSH level and pattern of mitochondrial distribution） and higher developmental potential after in vitro maturation （IVM） than the BCB-oocytes. Adult mice produced more BCB＋ oocytes with higher competence than the prepubertal mice when not primed with PMSG. PMSG priming increased both proportion and developmental potency of BCB＋ oocytes. The BCB＋ oocytes in the large-size group showed more SN chromatin configurations, better cytoplasmic maturity and higher developmental potential than their counterparts in the medium-size group. It is concluded that BCB staining can be used as an efficient method for oocyte selection, but that the competence of the BCB＋ oocytes may vary with oocyte diameter, animal sexual maturity and gonadotropin stimulation. Taken together, the series of criteria described here would allow for better choices in selecting oocytes for better development.

Effect of zeaxanthin on porcine embryonic development during in vitro maturation

Zeaxanthin is a common carotenoid, which is a powerful antioxidant that protects against damage caused by reactive oxygen species. The aim of the present study was to investigate the effects of zeaxanthin supplementation on in vitro maturation of porcine embryo development. We investigated nuclear maturation, intracellular glutathione （GSH）, and reactive oxygen species （ROS） levels during in vitro maturation, and subsequent embryonic development following parthenogenetic activation and in vitro fertilization OVF）. The oocytes were maturated and used at the metaphase II stage. After 42 hours of in vitro maturation, the zeaxanthin-treated group （0.5 μmol/L） showed significant increases in nuclear maturation （89.6%） than the control group （83.4%） （P 〈 0.05）. The intracellular GSH levels increased significantly （P 〈 0.05） as zeaxanthin concentrations increased; ROS generation levels decreased with increased zeaxanthin concentrations, but there were no significant differences. There were no significant differences in subsequent embryonic development, cleavage rate, blastocyst stage rate, and total blastocyst cell numbers following parthenogenetic activation and IVF when in vitro maturation media was supplemented with zeaxanthin. These results suggest that treatment with zeaxanthin during in vitro maturation improved the nuclear maturation of porcine oocytes by increasing the intracellular GSH level, thereby slightly decreasing the intracellular ROS level.

The Role of Brain-derived Neurotrophic Factor in Mouse Oocyte Maturation in vitro

Brain-derived neurotrophic factor (BDNF) can promote developmental competence in mammalian oocytes during in vitro maturation (IVM),but the role of BDNF in oocyte maturation at cellular level is not still clear.In this study,mouse cumulus-enclosed oocytes subjected to IVM were fertilized and cultured to blastocyst stage.Meiotic spindle configuration and cortical granules distribution during oocyte maturation in vitro were assessed by using immunofluorescence and laser confocal microscopy.The results showed that BDNF contributed to the complete preimplantation development of mouse oocytes compared to the control oocytes (13.78% vs.5.92%;P【0.05).Further,BDNF did not accelerate nuclear maturation of IVM oocytes.For the BDNF-treated oocytes at meiosis Ⅰ,Meiotic spindle areas were significantly smaller and the number of cytoplasmic microtubule organizing centers was greater than that in the control,and the percentages of oocytes showed spindles positioned near the oolemma and a well-formed cortical granule-free domain were significantly higher than that of the control.These morphological characteristics of the BDNF-treated oocytes were much closer to the oocytes matured in vivo than those of the control oocytes.In conclusion,BDNF can promote the developmental competence of mouse IVM oocytes,by improving the meiotic spindle configuration and location and cortical granules distribution at meiosis Ⅰ.

Resveratrol compares with melatonin in improving in vitro porcine oocyte maturation under heat stress

Background： Resveratrol, an important phyto-antioxidant commonly found in grapes, mulberry, and other plants,has a variety of functions including anti-aging, anti-cancer and anti-inflammatory activities. In the current study, we investigated the beneficial effects of resveratrol on in vitro porcine oocyte maturation under heat stress（HS）. The effect of resveratrol, melatonin and their combination on alleviating HS was compared according to the maturation rate of oocytes and the development competence of embryos after parthenogenetic activation（PA）.Results： Supplementation with resveratrol（2.0 μmol/L） not only improved the nuclear maturation but also raised the blastocyst rate of porcine embryos＇ PA from oocytes that underwent HS by increasing their glutathione（GSH）level, reducing reactive oxygen species（ROS） and up-regulating the expression of Sirtuin 1（SIRT1）. It was also found that melatonin（10^-7mol/L） and the combination of resveratrol（2.0 μmol/L） plus melatonin（10^-7mol/L） exhibited more potent effects than resveratrol alone regarding their protective activities on oocyte maturation under HS.Conclusions： This study compared the efficiencies of resveratrol, melatonin and their combination for protecting porcine oocytes from heat stress. The mechanisms are attributed to the fact that each treatment may have different ability to regulate the synthesis of steroid hormones and the expression of mature related genes.

Probiotic metabolites from Bacillus coagulans GanedenBC30^(TM) support maturation of antigen-presenting cells in vitro

AIM:To study the effects of probiotic metabolites on maturation stage of antigen-presenting immune cells.METHODS:Ganeden Bacillus coagulans 30(GBC30) bacterial cultures in log phase were used to isolate the secreted metabolite(MET) fraction.A second fraction was made to generate a crude cell-wall-enriched fraction,by centrifugation and lysis,followed by washing.A preparation of MET was subjected to size exclusion centrifugation,generating three fractions:< 3 kDa,3-30 kDa,and 30-200 kDa and activities were tested in comparison to crude MET and cell wall in primary cultures of human peripheral blood mononuclear cell(PBMC) as a source of antigen-presenting mononuclear phagocytes.The maturation status of mononuclear phagocytes was evaluated by staining with monoclonal antibodies towards CD14,CD16,CD80 and CD86 and analyzed by flow cytometry.RESULTS:Treatment of PBMC with MET supported maturation of mononuclear phagocytes toward both macrophage and dendritic cell phenotypes.The biological activity unique to the metabolites included a reduction of CD14+ CD16+ pro-inflammatory cells,and this property was associated with the high molecular weight metabolite fraction.Changes were also seen for the dendritic cell maturation markers CD80 and CD86.On CD14dim cells,an increase in both CD80 and CD86 expression was seen,in contrast to a selective increase in CD86 expression on CD14bright cells.The co-expression of CD80 and CD86 indicates effective antigen presentation to T cells and support of T helper cell differentiation.The selective expression of CD86 in the absence of CD80 points to a role in generating T regulatory cells.CONCLUSION:The data show that a primary mechanism of action of GBC30 metabolites involves support of more mature phenotypes of antigen-presenting cells,important for immunological decision-making.

Effects of Ovarian Preservation Time on in vitro Maturation and in vitro Fertilization of Oocytes from Slaughtered Sheep

[Objectives] The aim was to study the effects of ovarian preservation time on in vitro fertilization of oocytes from slaughtered sheep. [Methods] The collected ovaries were randomly and evenly divided into four groups. They were preserved in physiological saline containing penicillin（ 100 IU/ml） and streptomycin（ 100 μg/ml） at 15-20 ℃ for 0（ Control）,6,12 and 18 h,respectively. Then,the oocytes were subjected to in vitro fertilization. [Results]The maturation rates,cleavage rates and blastocyst rates of the oocytes preserved for 6 and 12 h showed no significant differences compared with those of the oocytes preserved for0 h（ 72. 03%,70. 87% vs. 73. 68%; 74. 12%,72. 60% vs. 74. 49%; 22. 22%,20. 75% vs. 23. 29%）（ P 〉 0. 05）. There were also no significant differences in maturation rate,cleavage rate or blastocyst rate between the oocytes preserved for 18 and 0 h（ P 〉 0. 05）. [Conclusions] Within a certain rage（ 0-18 h）,storage time of ovary at 15-20 ℃ does not affect the continued development of oocytes from slaughtered sheep.

Oxygen tension modulates cell function in an in vitro three-dimensional glioblastoma tumor model

Hypoxia is a typical feature of the tumor microenvironment,one of the most critical factors affecting cell behavior and tumor progression.However,the lack of tumor models able to precisely emulate natural brain tumor tissue has impeded the study of the effects of hypoxia on the progression and growth of tumor cells.This study reports a three-dimensional(3D)brain tumor model obtained by encapsulating U87MG(U87)cells in a hydrogel containing type I collagen.It also documents the effect of various oxygen concentrations(1%,7%,and 21%)in the culture environment on U87 cell morphology,proliferation,viability,cell cycle,apoptosis rate,and migration.Finally,it compares two-dimensional(2D)and 3D cultures.For comparison purposes,cells cultured in flat culture dishes were used as the control(2D model).Cells cultured in the 3D model proliferated more slowly but had a higher apoptosis rate and proportion of cells in the resting phase(G0 phase)/gap I phase(G1 phase)than those cultured in the 2D model.Besides,the two models yielded significantly different cell morphologies.Finally,hypoxia(e.g.,1%O2)affected cell morphology,slowed cell growth,reduced cell viability,and increased the apoptosis rate in the 3D model.These results indicate that the constructed 3D model is effective for investigating the effects of biological and chemical factors on cell morphology and function,and can be more representative of the tumor microenvironment than 2D culture systems.The developed 3D glioblastoma tumor model is equally applicable to other studies in pharmacology and pathology.

Adsorption,in vitro digestion and human gut microbiota regulation characteristics of three Poria cocos polysaccharides

Poria cocos(PC)is a famous traditional Chinese medicine(TCM)and a widely used healthcare ingredient,which has antiobesity,enhancing immunity and improving sleep effects.Traditionally,only water-soluble poria polysaccharide(WSP)is extracted and applied for clinical application,while insoluble polysaccharide(alkali-soluble poria polysaccharide,ASP)is discarded as herb residue.However,the whole PC has also been historically utilized as functional herbal food.Considering the beneficial role of dietary fiber and the traditional use of PC,ASP may also contribute substantially to the therapy function of PC.Compared to WSP,little attention has been paid to ASP and ASP modified product carboxymethyl poria polysaccharide(CMP)which has been used as an antitumor adjuvant drug.In this study,the oil,cholesterol,metal ions and polyphenols adsorption ability,in vitro simulated digestive and the gut microbiota fermentation characteristics of WSP,ASP and CMP were studied to evaluate the functional values of three P.cocos polysaccharides(PCPs).The results showed that all three PCPs had good adsorption capacity on cholesterol,polyphenols and metal ions(Cd^(2+)/Zn^(2+)/Mg^(2+)),among which ASP showed the highest capacity than WSP and CMP.The adsorption capacity of all three PCPs on heavy metal ions(Cd^(2+)/Zn^(2+))was stronger than that of non-heavy metal ions(Mg^(2+));The in vitro digestibility of all three PCPs was very low,but WSP was slightly higher than ASP and CMP;Moreover,the indigestible residue of all three PCPs could improve the richness and diversity of gut microbiota,among which ASP had the greatest influence.In general,ASP and CMP could significantly promote the proliferation of some probiotics and inhibit the growth of some harmful bacteria.The gut microbiota diversity of CMP was reduced,but the richness of probiotics,especially Parabacteroides distasonis was significantly enhanced compared with the ASP group,and the growth of harmful bacteria Klebsiella pneumoniae was inhibited after CMP treatment.The short-chain fatty acids(SCFAs)analysis results showed that all three PCPs could significantly promote the production of acetic acid,propionic acid and the total acid content compared with blank control group,and SCFAs producing activity was positively correlated with the proliferative capacity of probiotics.Taken together,the good adsorption characteristics and gut microbiota regulatory activity of ASP may lay foundation for its lipid-lowering and immune-improving function.Additionally,the probiotic effect of CMP and ASP indicated that except for only use the water extract of PC in clinic,CMP and ASP also can be used in healthcare to take full advantage of this valuable medicine.

A Review of Main Factors Involved in the Maturation of Oil Palm (Elaeis guineensis Jacq.) Fruit Bunches

The oil palm (Elaeis guineensis Jacq.) is a diploid perennial plant of the Arecaceae family. It is the most important plant cultivated for oil production. To ensure this production, certain optimal conditions are required: temperature, sunshine, rainfall, etc. The oil palm ensures its survival through the fruits borne on bunches located at the axis of the 17th to 20th leaves from the central stem. From pollination to the maturity of a bunch it takes about 4.5 to 6 months. Several events occur during this period: seed enlargement, weight increase, colour change, etc., but also important physiological changes: synthesis of some pigments (anthocyanin), increase in oil content correlated with the decrease in water content, etc. All of these constitute factors that can provide a better understanding of the biology of the seed. The aim of this work was to review some of the important parameters involved in the development and maturation of oil palm fruit bunches. These factors are classified into physiological, biochemical as well as environmental. The physiological parameters are color, appearance of embryo, seed weight and fruit detachment from bunches;Biochemical parameters include water content, oil content, carbohydrate, protein, mineral contents and lipase activity while temperature is the main environmental factor that affects fruit maturation. Thorough research has not yet been done at the different stages of maturation and ripening, thus a deep look into this may open up new avenues for research on early germinated oil palm seed production prior to seed dormancy.