Proteo-transcriptomic profiles reveal key regulatory pathways and functions of LDHA in the ovulation of domestic chickens(Gallus gallus)

Background In poultry, the smooth transition of follicles from the preovulatory-to-postovulatory phase impacts egg production in hens and can benefit the poultry industry. However, the regulatory mechanism underlying follicular ovulation in avians is a complex biological process that remains unclear.Results Critical biochemical events involved in ovulation in domestic chickens(Gallus gallus) were evaluated by transcriptomics, proteomics, and in vitro assays. Comparative transcriptome analyses of the largest preovulatory follicle(F1) and postovulatory follicle(POF1) in continuous laying(CL) and intermittent laying(IL) chickens indicated the greatest difference between CL_F1 and IL_F1, with 950 differentially expressed genes(DEGs), and the smallest difference between CL_POF1 and IL_POF1, with 14 DEGs. Additionally, data-independent acquisition proteomics revealed 252 differentially abundant proteins between CL_F1 and IL_F1. Perivitelline membrane synthesis, steroid biosynthesis, lysosomes, and oxidative phosphorylation were identified as pivotal pathways contributing to ovulation regulation. In particular, the regulation of zona pellucida sperm-binding protein 3, plasminogen activator, cathepsin A, and lactate dehydrogenase A(LDHA) was shown to be essential for ovulation. Furthermore, the inhibition of LDHA decreased cell viability and promoted apoptosis of ovarian follicles in vitro.Conclusions This study reveals several important biochemical events involved in the process of ovulation, as well as crucial role of LDHA. These findings improve our understanding of ovulation and its regulatory mechanisms in avian species.

Usefulness of Newly Devised Clomiphene Citrate Administration Method Compared with the Conventional Method in Ovulation and Pregnancy

Objective: Since not all women wish to conceive a child through aggressive treatment, we investigated the usefulness of modified repeated intracyclic clomiphene citrate (CC) therapy (repeated CC therapy) as a newly devised administration method. Methods: We evaluated the effects of CC administration on menstrual cycle length and retrospectively compared ovulation and pregnancy in 220 women who received CC at our hospital. Patients in the conventional method group received 50 mg per day for five days, starting on the fifth day of menstruation (withdrawal bleeding). Groups with and without menstrual cycle shortening after conventional CC administration were compared. The repeated CC therapy group was also compared with the non-shortened group. Repeated CC therapy was administered for the first five days as in the conventional method, and a second five-day repeat treatment was administered after an interval of five to seven days. Pregnancy rates, including indirect pregnancies, were evaluated by three different methods. Results: Ovulation and pregnancy rates were significantly better in the shortened group than in the non-shortened group (P < 0.001 and P = 0.010, respectively). Even in the non-shortened group, ovulation and pregnancy rates including indirect pregnancies were significantly improved when ovulation was observed with repeated CC therapy (P < 0.001 and P = 0.022, respectively). Conclusions: For patients whose menstrual cycle was not improved or shortened, repeated CC therapy as the newly devised CC administration method is useful as the next step after the conventional CC administration method.

Clinical Study on the Therapeutic Effect of Combined Traditional Chinese Medicine with Acupoint Injection and Herbal Paste Application in Treating Ovulatory Dysfunction in Polycystic Ovary Syndrome (PCOS)

Objective: To observe the therapeutic effect of combined traditional Chinese medicine (TCM) therapy with acupoint injection and herbal patch application on patients with ovulatory dysfunction associated with polycystic ovary syndrome (PCOS). Methods: A total of 80 PCOS patients who received treatment at the First Affiliated Hospital of Guangxi University of Chinese Medicine between October 2022 and March 2024 were enrolled in this study. They were randomly divided into a control group (n = 40) and an observation group (n = 40). The control group received conventional Western medical treatments, while the observation group was administered TCM combined with acupoint injection and herbal patch application. Pre- and post-treatment conditions were compared between the two groups. Results: After treatment, the overall effective rate was significantly higher in the observation group compared to the control group (P 0.05). Following treatment, both ovulation and pregnancy rates increased in both groups, but the increase was more evident in the observation group, showing a statistically significant difference (P < 0.01). Conclusion: For patients with oligoovulation or anovulation due to PCOS, the combined use of TCM, acupoint injection, and herbal patch application appears to be effective in improving systemic symptoms and achieving notable clinical outcomes.

Ovule Development, Formation of Pollination Drop and Pollination Process in Taxus chinensis (Taxaceae)

Pollen morphology and hydration, ovule development, formation of pollination drop and the pollination process of Taxus chinensis (Pilg.) Rehd were investigated in detail. The results showed that mature pollen was one_celled, irregular in shape, nonsaccate, and the surface was covered with orbicules. When pollen became hydrated, the intine swelled and the exine bursted. The exine usually remained on the surface of water or on the pollination drop all the time, while the rest portion of the pollen sank into the drop of water or the pollination drop. In the late August, the down_curved female buds could be observed in the field trees. The downturning of the female buds was an characteristic feature discriminating from the vegetative buds. At this stage ovular primordium was present in the female bud with a dome_shaped nucellar primordium encircled by a rudimentary integument. The integument grew fast and overgrew the nucellus. In the March of the following year, the ovule formed a micropylar canal about 1?000 μm in length, 200-300 μm in width. Except the micropyle, the other part of the ovule was enclosed by 20-25 scales. During the early stage of the ovule development, the nucellar surface clearly showed cell configuration; but a membrane_like structure covering the nucellar apex appeared in the late stage of the ovule development, and subsequently broke down with nucellar cell disintegration before pollen shedding. The disintegrated cell products might partake in the formation of the pollination drop. Before and during pollination, the nucellar cells contained abundant mitochondria, endospermic reticulum and dictysomes with vesicles; the plasma membrane was undulated and there was no accumulation of electron_dense substances in the pericytoplasm. Pollination drops usually occurred at night, and disappeared by noon in the field tree. 15 days before pollination, the in vitro ovules from the field trees already had the capability of producing pollination drops, which could last for 4-5 days indoors. However, after these drops received pollen, they did not withdraw noticeably compared with the unpollinated ones. The retraction of the pollination drop was presumed not to be a nonmetabolic outcome but rather a result of evaporation.

Cone and Ovule Development in Platycladus orientalis (Cupressaceae)

Seed cone in Platycladus orientalis (L.) France consists of four or five pairs of decussate bracts. Usually, two pairs of the fertile bracts in the middle of the cone subtend six ovules, which initiate in an acropetal manner. Only one ovule presents on each of the upper fertile bract, while two ovules initiate from a common primordium in the axil of lower bracts. In Beijing, most female cones initiated in July. All parts of the cone formed before dormancy, which occurred during November to the next January. After pollination in March, bract morphology changed dramatically; intercalary growth of the bract base formed a conspicuous protuberance, in which inverted vascular system developed. Furthermore, ovules on different pairs of bracts initiated in an acropetal manner and two ovules in each lower fertile bract initiated from a common primordium, which was different from the basipetal initiation of ovules and independently formed single ovule as reported by Takaso in Calltris

Study on Relation between Endogenous Phytohormones and Ovule Abortion in Tetraploid Robinia pseudoacacia

This study was conducted to investigate ovule abortion of tetraploid Robinia pseudoacacia and its reasons. It was found that in the ovule development period of tetraploid R. pseudoacacia from 15 d after coronal opening, the levels of growth-promoting endogenous hormones in abortive ovules were lower than those in normal ovules, and the level of a growth-inhibiting hormone was always higher than those in normal ovules, indicating that ovule abortion of tetraploid R. pseudoacacia might be related to abnormal endogenous phytohormones in ovules. This study laid a foundation for further study on highly sterile phenomenon of tetraploid R. pseudoacacia seeds.

Comparative transcriptome analysis of the effect of different heat shock periods on the unfertilized ovule in watermelon(Citrullus lanatus)

In vitro gynogenesis is an important tool used in haploid or homozygous double-haploid plant breeding.However,because of low repeatability,embryoid induction rate and quality,the molecular mechanisms remain poorly understood.Heat shock treatment can promote the transformation of the gametophytic pathway into the sporophyte pathway,which induces the occurrence of haploid.In this study,unfertilized ovaries were heat shocked for 0 h(A0)before flowering and for 0 h(A1),4 h(A3),8 h(A5),12 h(A7),and 24 h(A8),respectively,at 37℃at the first day of the flowering stage.The ovule enlargement rate was increased from 0%at 25℃to 96.8%at 37℃(24 h treatment).Thus,we aimed to investigate the gene expression patterns in unfertilized ovules of watermelon after different periods of heat shock by using RNA-Seq technology.The results showed that compared with A3,A5,A7,and A8,the biosynthesis of amino acid,glycine,serine and threonine metabolic pathways in A1 has changed significantly.This indicated that heat shock treatment affected the synthesis and transformation of amino acids during ovule expansion.The transcriptome data suggested gene expressions of ovule growth were significantly changed by heat-specific influences.The results provide new information on the complex relationship between in vitro gynogenesis and temperature.This provides a basis for further study of the mechanism of heat shock affecting the expansion of watermelon ovule.

Interspecific Crossing between Yam Species (Dioscorea rotundata and Dioscorea bulbifera) through in Vitro Ovule Culture

In the present study, in vitro ovule culture technique was used to obtain interspecific cross combination of Dioscorea rotundata ufenyi and Dioscorea bulbifera wild. Ten days after pollination, ovules were excised and cultured onto 1/2 strength Murashige and Skoog (MS) medium (Basal salt mixture + Vitamins) supplemented with 6% sucrose, 0.7% agar and plant growth hormones such as GA3, BAP, Picrolam and TDZ. Cultured ovules were transferred on 1/2 MS medium with 3% sucrose and 0.7% agar after three weeks. 40 days after pollination, germination was observed from 7 months cultured ovule between D. rotundata ufenyi x D. bulbifera wild. Hybridity of the regenerated plant was checked by flow cytometric method. A close relation was observed between the fluorescence intensity of the obtained progeny with one of the parents’ fluorescence. The observed progeny can be closely correlated with an apomictic tissue from an ovule parent of D. rotundata ufenyi. Plantlets derived from ovule culture were proliferated through in vitro shoot multiplication with hormonal concentration (0.5 mg/l BAP) supplemented with 1/2 strength MS medium. Obtained ovule culture derived in vitro plantlets were successfully hardened, acclimatized and transferred to the field, where they survived and grew normally. In plant breeding, interspecific crossing is very important technique, enabling the time needed to produce homozygous lines to be shortened as compared to the conventional plant breeding techniques.

Ectopic expression of a male fertility gene,LOGL8,represses LOG and hinders panicle and ovule development

Grain number and seed-setting rate are components of crop yield.Cytokinin influences grain yield.However,emerging studies suggest that high cytokinin signals often lead to reduced branching or seed-setting rate,leading to reduced grain yield,although the mechanisms remain unclear.In this study,we identified and characterized the rice(Oryza sativa L.)gene LONELY GUY-LIKE 8(LOGL8),based on analysis of the LOGL8-pm(promoter mutant of LOGL8)mutant,which harbors a T-DNA insertion in the promoter of this gene.The mutation in LOGL8-pm causes ectopic hyperexpression of LOGL8 in inflorescence organs,resulting in plants with smaller panicles and defective ovules lacking archesporial cells and integuments.Knockout of LOGL8 caused pollen abortion,leading to a reduced seed-setting rate.LOGL8 encodes a putative cytokinin-activating enzyme.Our results showed that LOGL8 directly catalyzes the biosynthesis of bioactive cytokinins.Therefore,we propose that the ectopic expression of LOGL8 disrupts cytokinin spatiotemporal distribution and causes inhibition of LONELY GUY(LOG),which affects panicle branching and female organ development.These findings reveal the important role of LOGL8 in male development,and highlight the delicate balance of local cytokinin levels during panicle branching and female organ development.

New observation on seed/ovule position in the fruit of Archaeanthus and its systematic implications

1.Introduction Angiosperms are the most important group of plants in the current earth ecosystem.However,little is known about the origin of angiosperms,which has been a headache for many botanists.The core question in the origin of angiosperms is the homology of carpels.Formerly,Arber EAN and Parkin J(1907)speculated that the carpels in angiosperms were derived from the former megasporophylls bearing ovules along their margins.

Anatomical Study on the Multi-Ovule Development and Abortion of Hanfu Apple

The terminal flower buds of 6-yr-old Hanfu apple were used to study the ovule development, ovular characteristics, cell death of abortive ovules, and dynamic change of starch grain quantity in the embryo sac with paraffin slices and terminal deoxynucleotidyl transferase-mediated fluorescein deoxyuridine triphosphate nick-end labeling （TUNEL） system. Four anatropous ovules in each ventricle could be observed before flowering. With the developing of floral organ, the bulk of normal ovules enlarged in each ventricle, the mature embryo sac differentiated into nucellus, and the egg cell developed into zygote by double fertilization. A large number of starch grains were observed during pollen tube growth and double fertilization, which guaranteed basic nutrient supply in the normal development of ovules. Moreover, abortion phenomenon of runtish ovules emerged at the stages of mature embryo sac, double fertilization and zygote development. The abortion characteristics included deformity of ovule development, degradation of nucellus tissue, separation between funiculus and ovule, abnormality of four-nucleate embryo sac, as well as development interruption of mature embryo sac. TUNEL analysis proved that ovule abortion was programmed cell death.

Clinical Application of Double Ovulation Stimulation in Patients with Diminished Ovarian Reserve and Asynchronous Follicular Development Undergoing Assisted Reproduction Technology

Objective This study aimed to compare the clinical effects of double ovulation stimulation(DouStim)applied during the follicular and luteal phases with the antagonist protocol in patients with diminished ovarian reserve(DOR)and asynchronous follicular development undergoing assisted reproductive technology(ART).Methods The clinical data of patients with DOR and asynchronous follicular development receiving ART from January 2020 to December 2021 were retrospectively analyzed.The patients were divided into two groups according to their ovulation stimulation protocol:DouStim group(n=30)and antagonist group(n=62).Assisted reproduction and clinical pregnancy outcomes were compared between the two groups.Results In the DouStim group,the number of oocytes retrieved,metaphase II(MII)oocytes,two-pronuclei(2PN),day 3(D3)embryos,D3 high-quality embryos as well as blastocyst formation,implantation,and human chorionic gonadotropin-positive rates were significantly greater than those in the antagonist group(all P<0.05).No significant differences were found in MII,fertilization,or continued pregnancy rates at the first frozen embryo transfer(FET),in-vitro fertilization(IVF)cancellation,or early medical abortion rates between the groups(all P>0.05).Except for the early medical abortion rate,the DouStim group generally had favorable outcomes.In the DouStim group,the dosage and duration of gonadotropin and the fertilization rate were significantly greater in the first ovulation stimulation induction than in the second ovulation stimulation induction(P<0.05).Conclusion The DouStim protocol efficiently and economically obtained more mature oocytes and high-quality embryos for patients with DOR and asynchronous follicular development.

Global Annotation of Small RNA and MicroRNA Mature Sequences from Developing Ovules of Gossypium hirsutum L.

The involvement of small RNAs in cotton fiber development is under explored.The objective of this work was to directly clone,annotate,and analyze small RNAs of developing ovules to reveal

The sperm-interacting proteome in the bovine isthmus and ampulla during the periovulatory period

Background Spermatozoa interact with oviduct secretions before fertilization in vivo but the molecular players of this dialog and underlying dynamics remain largely unknown.Our objectives were to identify an exhaustive list of sperm-interacting proteins(SIPs)in the bovine oviduct fluid and to evaluate the impact of the oviduct anatomical region(isthmus vs.ampulla)and time relative to ovulation(pre-ovulatory vs.post-ovulatory)on SIPs number and abundance.Methods Pools of oviduct fluid(OF)from the pre-ovulatory ampulla,pre-ovulatory isthmus,post-ovulatory ampulla,and post-ovulatory isthmus in the side of ovulation were collected from the slaughterhouse.Frozen-thawed bull sperm were incubated with OF or phosphate-buffered saline(control)for 60 min at 38.5℃.After protein extraction and digestion,sperm and OF samples were analyzed by nanoLC-MS/MS and label-free protein quantification.Results A quantitative comparison between proteins identified in sperm and OF samples(2333 and 2471 proteins,respectively)allowed for the identification of 245 SIPs.The highest number(187)were found in the pre-ovulatory isthmus,i.e.,time and place of the sperm reservoir.In total,41 SIPs(17%)were differentially abundant between stages in a given region or between regions at a given stage and 76 SIPs(31%)were identified in only one region×stage condition.Functional analysis of SIPs predicted roles in cell response to stress,regulation of cell motility,fertilization,and early embryo development.Conclusion This study provides a comprehensive list of SIPs in the bovine oviduct and evidences dynamic spatiotemporal changes in sperm-oviduct interactions around ovulation time.Moreover,these data provide protein candidates to improve sperm conservation and in vitro fertilization media.

Research Progress on Mechanism of Ovulation Disorder in Polycystic Ovary Syndrome

Polycystic ovary syndrome(PCOS)is a reproductive endocrine disease characterized by ovulation disorder,hyperandrogenism and/or polycystic ovary.Follicular development disorder caused by PCOS accounts for up to 75%of anovulatory infertility,which is a common cause of infertility and affects up to 15%of reproductive aged women worldwide.Oxidative stress,chronic inflammation,endoplasmic reticulum stress and other pathological conditions coexist in the ovarian microenvironment of PCOS,which are further aggravated under the action of high levels of androgens,synergistically deteriorating the follicular microenvironment,leading to ovulation disorders in PCOS patients.This paper briefly reviewed the research on the mechanism of PCOS ovulation disorder in recent years.

三促汤联合绒促性素对FM/TE不孕症患者排卵率、妊娠率的影响及对临床疗效的探究

[目的]探究三促汤联合绒促性素对卵泡发育不良(FM)/薄型子宫内膜(TE)不孕症患者排卵率、妊娠率的影响及对临床的疗效。[方法]选取广西中医药大学附属瑞康医院2022年7—10月于门诊就诊的105例不孕症患者为研究对象,随机数字表法将105例患者分为35例中药组,35例西药组和35例试验组。中药组:促卵泡汤+促排卵汤+促黄体汤;西药组:克罗米芬(CC)+绒促性素(HCG)2 000 IU肌肉注射;试验组:(促卵泡汤+CC)+(促排卵汤+HCG 2 000 IU肌肉注射)+促黄体汤。以此检测各组患者排卵率、妊娠率、卵泡的最大直径、子宫内膜的厚度、血液流变学指标、卵泡刺激素(FSH)、促黄体生成素(LH)、雌二醇(E2)水平、中医证候评分、临床疗效的情况变化。[结果]与中药组、西药组比较,试验组患者排卵率、妊娠率、临床疗效升高,不良反应发生率较低(P<0.05)。与治疗前比较,治疗后各组患者卵泡的最大直径、子宫内膜的厚度、FSH、E2水平升高,血沉、全血低切黏度、血浆黏度、全血高切黏度、胸闷胀痛、月经不调、心烦不舒、易怒烦躁、小腹胀痛、乳房胀痛评分降低(P<0.05),与中药组、西药组治疗后比较,试验组患者治疗后卵泡的最大直径、子宫内膜的厚度、FSH、E2有所上升(P<0.05),治疗后试验组患者血沉、全血低切黏度、血浆黏度、全血高切黏度、胸闷胀痛、月经不调、心烦不舒、易怒烦躁、小腹胀痛、乳房胀痛评分及总积分降低(P<0.05),而LH表达差异无统计学意义(P>0.05)。[结论]通过使用三促汤联合HCG治疗,可有效改善患者妊娠率及排卵率,调节机体性激素及血液流变学相关指标水平,提高临床疗效。

PPOS促排卵方案对颗粒细胞芳香化酶CYP19A1和ERβ受体表达的自身对照研究

目的研究第一周期行黄体期减量长方案行体外受精-胚胎移植(IVF-ET)促排卵过程中黄体生成素(LH)偏低(扳机日LH<1.2 mIU/mL)的反复种植失败(RIF)患者,第2周期改用高孕激素状态下促排卵(PPOS)方案。检测两种促排卵方案中颗粒细胞中芳香化酶CYP19A1和ERβ受体的mRNA和蛋白质表达水平,以探讨不同促排卵方案对卵泡颗粒细胞发育的影响。方法对2020年1月至2022年1月在南通大学附属南通妇幼保健院生殖医学中心行IVF-ET的RIF患者进行研究,第1周期采用黄体期减量长方案RIF患者,第2周期改采用PPOS方案患者共36例。同时收集两种方案促排卵卵泡液中壁层颗粒细胞,检测颗粒细胞中芳香化酶CYP19A1和ERβ受体的mRNA和蛋白质表达水平,自身对照分析两种促排卵方案的临床和实验室指标,研究不同促排卵方案对卵泡颗粒细胞发育的影响。结果PPOS方案和长方案平均获卵数、成熟卵母细胞数的比较差异无统计学意义(P>0.05),而PPOS方案的扳机日LH水平、正常受精(2PN)数、正常受精率、优质胚胎率显著高于长方案(P<0.05)。PPOS组中CYP19A1和ERβ受体的mRNA表达水平均高于对照组,差异有统计学意义(P<0.001)。与mRNA表达水平结果相一致,PPOS组中CYP19A1和ERβ受体的蛋白表达水平均高于对照组,差异有统计学意义(P<0.05)。结论第1周期行长方案促排卵过程中LH偏低的RIF患者中,第2周期改用PPOS方案组后,扳机日的LH水平明显上升,同时壁层颗粒细胞ERβmRNA水平和蛋白水平均显著高于长方案组,CYP19A1 mRNA水平和蛋白水平均也显著高于长方案组,改用PPOS方案后,LH水平有所提高,改善了患者的卵子质量,进而提高了胚胎质量。

西曲瑞克联合阿司匹林预防早发型卵巢过度刺激综合征的疗效观察

目的观察西曲瑞克联合阿司匹林预防早发型卵巢过度刺激综合征(OHSS)的疗效。方法回顾性收集2022年1月1日至7月1日于我院治疗的38例早发型OHSS患者资料,根据用药方案的不同分为对照组(19例)和干预组(19例)。取卵术后第1天,对照组患者给予阿司匹林肠溶片100 mg,口服至月经来潮;干预组患者给予注射用西曲瑞克0.25 mg,连续3 d皮下注射+阿司匹林肠溶片(用法用量同对照组)。观察并检测两组患者的首个黄体期、OHSS严重程度分级及取卵术后卵巢体积、腹水体积、血清雌二醇(E_(2))、白细胞计数(WBC)、红细胞比容(HCT)、中性粒细胞百分比(NEUT%)、D-二聚体(DD)、凝血酶原时间(PT)、纤维蛋白原(Fib)。结果干预组患者的首个黄体期显著短于对照组,中、重度患者比例显著低于对照组(P<0.05或P<0.01)。取卵术后,干预组患者的卵巢体积、腹水体积、血清E_(2)、WBC、NEUT%、HCT、DD、Fib总体低于对照组,PT显著高于对照组(P<0.05)。结论西曲瑞克联合阿司匹林预防早发型OHSS的疗效较单用阿司匹林更好。

牛蒡繁育系统与花部特征研究

[目的]了解牛蒡生殖特性,为其栽培育种提供理论基础。[方法]通过野外观察,运用异交指数(OCI)、花粉胚珠比(P/O)及套袋试验等研究方法对大连地区辽宁中医药大学内牛蒡的花部特征及繁育系统进行研究。[结果]牛蒡群体花期一般为34~35 d,花期重叠,每年6月下旬开始到7月末结束,其花粉和柱头具有活力且柱头在全天都具有活力;OCI为4,P/O为5 666,异花授粉需要传粉者,风对其传粉作用不明显;套袋试验中自然套袋与自然对照均可结实,结实率分别为99.0%和69.5%,去雄去柱头后套袋结实率为0,不具有无融合生殖特性。[结论]牛蒡具有异交特性且自花授粉可结实,风媒传粉可能性小。

排卵障碍性不孕中医外治法研究进展

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