Proteome changes of porcine follicular fluid during follicle development

Background:Ovarian follicular fluid influences follicle and oocyte growth,but the fluctuation of its protein content during folliculogenesis has not been comprehensively analyzed.Here we used a shotgun approach and bioinformatics analyses to investigate and compare the proteomes of porcine follicular fluid(pFF)obtained from small(<4 mm),medium(4–6 mm)and large(>6–12 mm)follicles.Results:Follicular fluid samples containing highest estrogen levels were selected as non-atretic from small(SNA:26.1±15 ng/mL),medium(MNA:162±54 ng/mL),and large(LNA:290±37 ng/mL)follicles for proteomic analyses.We detected 1627,1699,and 1756 proteins in SNA,MNA,and LNA samples,respectively.Nearly 60–63%of total proteins were specific to each sample,11–13%were shared in pairwise comparisons,and 247 proteins were shared among all samples.Functional categorization indicated comparable gene ontology(GO)terms distribution per cellular component,molecular function,and biological process categories across samples;however,the ranking of highly significantly enriched GO terms per category revealed differences between samples.The patterns of proteinto-protein interactions varied throughout follicle development,and proteins such as serine protease inhibitor,clade E(SERPINE);plasminogen activator,urokinase(PLAU);and plasminogen activator,urokinase receptor(PLAUR)appeared stage-specific to SNA,MNA,and LNA,respectively.The“complement and coagulation cascades”was the common major pathway.Besides,properdin and fibulin-1 were abundant proteins that appeared absent in LNA samples.Conclusion:This study provides extensive and functional analyses of the pFF proteome changes during folliculogenesis and offers the potential for novel biomarker discovery in pFF for oocyte quality assessment.

Influence of Follicular Fluid on <i>in Vitro</i>Maturation and Fertilization of Bovine Oocytes

The aim of this study was to investigate the effect of time on in vitro maturation of bovine oocytes and of the addition of follicular fluid on meiotic progression. The cumulus-oocyte complexes (COCs) collected from 3 to 6 mm follicles were obtained from ovaries of slaughtered female animals. The medium of maturation was supplemented or not with 20 μL follicular fluid (FF);661 oocytes were matured in vitro (extrusion of the first polar corpuscle) for 22 hours with added follicular fluid (AFF) (72.01%) or without follicular fluid (WFF) (67.53%) and 679 oocytes were matured in vitro for 26 hours (extrusion of the first polar corpuscle) with AFF (92.1%) and WFF (77.15%). The results of extrusion of the second polar corpuscle as an event related to the fertilization percentages showed that the increase in the fertilization rate is maintained at 26 hours with AFF (79.45%), but the percentage decreases WFF (65.08%). After 22 hours, the fertilization rate was 62.38% AFF and 53.40% WFF. The developmental competence of bovine oocytes is affected by the duration of maturation in vitro and the inclusion in the FF culture medium. The use of follicular fluid in the in vitro maturation medium may be a biological strategy to increase the cumulus expansion, the nuclear maturation and the in vitro fertilization.

Association of the microbial culture of follicular fluid,vaginal swab and catheter tip with β-hCG IVF positive and negative

Objective:To find out the association of microbial contamination withβ-human chorionic gonadotropin(β-hCG)in-vitro fertilization(IVF)positive and negative.Methods:A total of 73 fresh IVF cycle women were included in the retrospective study.Vaginal swab culture samples were collected prior to ovum pick-up and embryo transfer.The follicular fluids were collected during ovum pick-up and catheter tip culture samples were collected after successful embryo transfer.After 14 days of the embryo transfer,women were classified intoβ-hCG IVF positive and negative.The comparative statistical analyses of aerobic microbial culture reports were done betweenβ-hCG IVF positive and negative women.Results:Out of 73 women,42(57.5%)were found to beβ-hCG IVF positive and 31(43.5%)were negative.In the aerobic culture of ovum pick-up vaginal swab,follicular fluid,embryo transplantation vaginal swab and catheter tip,Enterococcus faecalis was found to be higher compared to other bacteria(Streptococcus spp.,Candida,Escherichia coli and Klebsiella).Regarding the comparison between IVF positive and negative,the overall microbial infection rate of vaginal swab culture during ovum pick-up and embryo transplantation was found to be higher in IVF negative women than in IVF positive women(38.71%vs.28.57%);however,it was not statistically significant(P>0.05).The follicular fluid and catheter tip culture microbial infection rate was found to slightly higher in IVF positive women than in IVF negative women(54.76%vs.41.94%and 19.05%vs.9.68%,respectively),but there were not significant differences(P>0.05).Conclusions:The aerobic microbial culture reports of follicular fluid,vaginal swab culture,and catheter tip culture are not statistically significantly withβ-hCG IVF positive.

Protein Expression of Goat Follicular Fluid （GFF）

Oocyte maturation process is very complex. Until now synthesis and protein function as well as pathway mechanism in oocyte maturation process are not yet well understood. The mechanism of goat follicular fluid （GFF） in maturation in vitro medium needs to be deeply studied before it is implemented widely. The aim of the research was analyzing the GFF to know protein expression of ERK1, ERK 2, p90rsk, Cyclin-Bl and cdc25A by Western Blotting method. GFF collection by aspiration from small follicle （〈 4 mm） and big follicle （4-8 mm） by using needle with spuit size of 18 G. It was concluded that there was protein expression of ERK2 and p90rsk, at 42 kDa and 22 kDa respectively.

Computer Simulation of Gd(III) Speciation in Human Interstitial Fluid

The speciation and distribution of Gd(III) in human interstitial fluid was studied by computer simulation. Meantime artificial neural network was applied to the estimation of log ?values of complexes. The results show that the precipitate species, GdPO4 and Gd2(CO3)3, are the predominant species. Among soluble species, the free Gd(III), [Gd(HSA)] , [Gd(Ox)] and then the ternary complexes of Gd(III) with citrate are main species and [Gd3(OH)4] becomes the predominant species at the Gd(III) total concentration of 2.2?0-2mol/L.

Computer Simulation of Insoluble Pr(III) Speciation in Human Interstitial Fluid

A multi-phase model of Pr(III) speciation in human interstitial fluid was constructed and insoluble Pr(III) speciation was studied. When the total concentration of Pr(III) is below 8.401E-10 mol/L, soluble Pr(III) species are main species. With rising the total concentration of Pr(III), Pr(III) is firstly bound to phosphate to form precipitate of PrPO4, then bound to carbonate and another precipitate of Pr2(CO3)3 was obtained. When the total concentration is between 1.583E-9 mol/L and 4.000E-3 mol/L, the insoluble species are predominant Pr(III) species.

MATHEMATICAL STUDY OF DIFFUSIVE FLUID TRANSPORT AND DISTRIBUTION IN HUMAN DERMAL REGIONS

This paper deals with mathematical study of diffusive fluid transport and distri- bution in human dermal parts. It accounts the intracellular fluid which continuously flows through the skin shells in order to maintain fluid balance within the body. A mathematical model is envisaged for this process and the finite element method （FEM） is employed to calculate the concentration of the fluid at different skin layers. This estimation is analyzed in relation with other parameters of the tissue medium and the atmosphere.

Follicular fluid composition of ovulatory follicles in repeat breeder Holstein dairy cows

Objective: To examine differences in the metabolite, steroid and lipopolysaccharide of follicular fluid collected from the ovulatory follicle of Holstein repeat breeding cows, lactating cows, and virgin fertile heifers. Methods: Estrus was induced in animals possessing functional corpus luteum by intramuscular administration of prostaglandin F2α. Six to twelve hours after detection of the standing estrus, cervical samples were collected to detect subclinical endometritis via counting neutrophils on stained smears of cervical swabs. Then, follicular fluid of ovulatory follicles and serum samples were collected from repeat breeding cows (n=11), lactating cows (n=8) and virgin fertile heifers (n=10). Sodium and potassium were measured with a flame photometer method. Urea, total protein, glucose, cholesterol andβ-hydroxybutyric acid were assayed with commercial spectrophotometry kits. Chloride concentration was also measured with titration of samples against silver nitrate. Progesterone, estradiol-17βand lipopolysaccharide concentrations were measured using enzyme-linked immuno sorbent assay kits. Results: All analysis of follicular fluid samples showed that repeat breeding and lactating cows had a mean higher lipopolysaccharide concentration than that of the virgin fertile heifers (P<0.05). But concentration of serum estradiol-17βin repeat breeding and lactating cows was lower than that of virgin fertile heifers (P<0.05). In addition, the mean percentage of neutrophils in the cervical secretion of repeat breeding cows was higher than that of lactating cows (P<0.05). Conclusions: High follicular fluid concentration of lipopolysaccharide in ovulatory follicles results in the occurrence of repeat breeding syndrome in dairy cows. Further, a lower serum estradiol-17βconcentration and a higher percentage of neutrophil in the cervical secretion on the day of artificial insemination may have resulted in the occurrence of repeat breeding syndrome in dairy cows.

Effect of Follicular Fluid Lactoferrin Level on Oocytes Quality and Pregnancy Rate in Intracytoplasmic Sperm Injection Cycles

Assessment of oocyte quality to avoid overproduction of embryos is now considered an important goal in ICSI cycles. Although in classic Intra Cytoplasmic Sperm Injection (ICSI) cycles selection is mainly done for embryos and not for follicles. Follicular fluid and its contents representing the oocyte environment are now gaining more attention because not only of its crucial influence on oocyte developments but also due to easy isolation of follicular fluid with every case of ICSI. By its aspiration during ovum pick up (OPU). substances isolated from follicular fluid to assess oocyte quality were in the form of: hormones like, Follicle Stimulating Hormone (FSH), Luteinizing Hormone (LH), Anti-mullerian Hormon (AMH), Growth Hormone (GH) etc., growth factors like Insulin Like Growth Factor (ILGF), proteins and amino acids like Lactoferrin (LF) which is an iron-binding glycoprotein that was detected in follicular fluid and was thought to be related to good oocyte quality when present in high concentration. Objectives: The aim of this study was to detect the possible effect of follicular fluid lactoferrin on oocyte quality and hence pregnancy rate in ICSI cycles. Methods: Follicular fluid was obtained from 64 patients undergoing Intra-cytoplasmic sperm injection (ICSI) procedure in Ain Shams University Maternity Hospital. Follicular fluid (FF) was collected at the time of oocyte harvesting. The lactoferrin concentration in FF was assayed by ELISA. Results: The mean LF concentration in follicular fluid of the positive biochemical pregnancy group (0.63 ± 0.17 ng/mL) was not significantly higher than that in the negative biochemical pregnancy (0.61 ± 0.16 ng/mL). A positive correlation between a number of mature oocytes and lactoferrin concentration was not found (r.101). Conclusion: Lactoferrin level in follicular fluid does not correlate with oocytes quality or pregnancy rate.

The Analysis of Human Behaviour and Evacuation in a Fire Situation in a Building for Higher Education

The paper presents the analysis of a human evacuation from a higher education building located in Iassy,Romania,by means of engineering techniques to approach fire safety.Because in Romania(as in other European countries)fire safety design of buildings is prescriptive and not performance-based,a fire safety engineering approach arouse great interest in many countries such as the U.S.A.,Australia,New Zealand,England,Sweden,Finland,etc.This paper is based on the assumption of starting a fire in the space of a hall for festivities,located on the ground floor of the building,near two human evacuation routes;We consider two building evacuation scenarios:two exits and,respectively,just one exit(assuming that the second would be accidentally blocked).

Seasonal influence on miRNA expression dynamics of extracellular vesicles in equine follicular fluid

Background Ovarian follicular fluid(FF)is a dynamic environment that changes with the seasons,affecting follicle development,ovulation,and oocyte quality.Cells in the follicles release tiny particles called extracellular vesicles(EVs)containing vital regulatory molecules,such as microRNAs(miRNAs).These miRNAs are pivotal in facilitating commu-nication within the follicles through diverse signaling and information transfer forms.EV-coupled miRNA signaling is implicated to be associated with ovarian function,follicle and oocyte growth and response to various environmen-tal insults.Herein,we investigated how seasonal variations directly influence the ovulatory and anovulatory states of ovarian follicles and how are they associated with follicular fluid EV-coupled miRNA dynamics in horses.Results Ultrasonographic monitoring and follicular fluid aspiration of preovulatory follicles in horses during the ano-vulatory(spring:non-breeding)and ovulatory(spring,summer,and fall:breeding)seasons and subsequent EV isola-tion and miRNA profiling identified significant variation in EV-miRNA cargo content.We identified 97 miRNAs with dif-ferential expression among the groups and specific clusters of miRNAs involved in the spring transition(miR-149,-200b,-206,-221,-328,and-615)and peak breeding period(including miR-143,-192,-451,-302b,-100,and let-7c).Bioinformatic analyses showed enrichments in various biological functions,e.g.,transcription factor activity,transcrip-tion and transcription regulation,nucleic acid binding,sequence-specific DNA binding,p53 signaling,and post-trans-lational modifications.Cluster analyses revealed distinct sets of significantly up-and down-regulated miRNAs associ-ated with spring anovulatory(Cluster 1)and summer ovulation–the peak breeding season(Clusters 4 and 6).Conclusions The findings from the current study shed light on the dynamics of FF-EV-coupled miRNAs in relation to equine ovulatory and anovulatory seasons,and their roles in understanding the mechanisms involved in seasonal shifts and ovulation during the breeding season warrant further investigation.

Molecular cloning and characterization of human age-related NADH oxidase (arNOX) proteins as members of the TM9 superfamily of transmembrane proteins

Age-related NADH oxidase (arNOX = ENOX3) proteins are superoxide-generating cell surface oxidases that increase in activity with age beginning at about 30 y. A soluble and truncated exfoliated form of the activity is present in blood and other body fluids. The activity was purified to apparent homogeneity from human urine and resolved by 2-D gel electrophoresis into a series of 24 to 32 kDa components of low isoelectric point. The purified proteins were resistant both to N-terminal sequencing and trypsin cleavage. Cleavage with pepsin revealed peptides corresponding to the TM9 family of transmembrane proteins. Peptide antisera raised to all five members of the human TM9 family sequentially blocked the arNOX activity of human saliva and sera. The soluble truncated N-terminus of the human homolog TM9SF4 was expressed in bacteria. The recombinant protein was characterized biochemically and exhibited ar-NOX activity. The findings identify five arNOX isoforms each of which correspond to one of the five known TM9 family members. The exfoliated soluble arNOX forms are derived from the 24 to 32 kDa N-termini exposed to the cell’s exterior at the cell surface. Each of the shed forms contain putative functional motifs characteristic of ECTO-NOX (ENOX) proteins despite only minimal sequence identity. Our findings identify arNOX as having functional characteristics of ENOX proteins and the TM9 superfamily of proteins as the genetic origins of the five known arNOX isoforms present in human sera, plasma and other body fluids1.

3D CFD Simulation of Pulsatile Blood Flow in the Human Aorta

A three-dimensional computational fluid dynamics (CFD) simulation of the physiological pulsatile blood flow in the human aortic arch and its three branches has been conducted by using commercial software StarCD. The blood flow, of a peak Reynolds number of 3289 and a Womersley parameter of 16.44, was simulated in a rigid aorta geometry that was built by computer aided design (CAD) reconstruction method based on autopsy data of a female adult. The purpose of this work is to further the understanding of the complex nature of aorta flow, therefore it mainly focuses on analysis of the spatial and temporal distributions of velocities and wall shear stresses. The results, illustrated by 3D visualization pictures and 2D graphs of the primary velocity profiles, wall shear stress and pressure distributions, as well as the secondary flow patterns, are in good agreement with those of other experimental and computational works. The distributions of pressure and wall shear stress support the correlation between high and low shear stresses and pressures and the atherosclerotic lesions.

Serodiagnosis of human hydatidosis with an ELISA developed based on antigens derived from sheep hydatid cysts and comparison with a commercial human ELISA kit

Objective:To explore the serodiagnosis of hydatid cyst in human using different antigens of sheep(hydatid fluid,somatic and Lxcretory/secretory antigens of protoscolexi by ELISA and compares this result with commercial human ELISA kit.Methods:one hundred blood samples from patients with history of severe abdominal pain and cosinophilia were obtained.Ten serum samples were obtained from surgically and pathologically confirmed cystic echinococcosis patients from Mashhad university hospital as positive control and 5 serum samples from infant under one year old as negative control.Blood samples were centrifuged at 3000xg at 20℃for 15 min and sera were stored at-20℃.First,these samples were tested for the presence of antibody by commercial human ELISA.Then.ELISA was developed on microplates coated with hydatid fluid,Somatic and Excretory/secretory antigens of protoscolex of sheep.Results:The results of this study as analyzed by Kappa test showed that,hydatid fluid antigen could be used as a precise source of detection in indirect ELISA test.Conclusions:Hydatid fluid in comparison with Excretory-secretory and somatic antigens showed more compatibility agreement in kappa test which can be used for further studies in development of any ELISA test for diagnosis of human hydatidosis.

Impact of human motion on TVOCs inhalation dose under side re-circulated ventilation

The main objectives were to (1) calculate the total volatile organic compounds (TVOCs) inhalation dose, (2) analyze the proportions of human’s inhaled contaminant dose from different sources, and (3) present a newly defined ratio of relative inhalation dose level (RIDL) to assess indoor air quality (IAQ). A user defined function based on CFD (computational fluid dynamics) was developed, which integrated human motion model with TVOCs emission model in a high sidewall air supply ventilation mode. Based on simulation results of 10 cases, it is shown that the spatial concentration distribution of TVOCs is affected by human motion. TVOCs diffusion characteristic of building material is the most effective way to impact the TVOCs inhalation dose. From the RIDL index, case A-2 has the most serious IAQ problem, while case D-1 is of the best IAQ.

Follicular fluid levels of prostaglandin E2 and the effect of prostaglandin E2 on steroidogenesis in granulosa-lutein cells in women with moderate and severe endometriosis undergoing in vitro fertilization and embryo transfer

Background The mechanisms of endometriosis with infertility have not been fully studied. The present study aimed to assess the follicular fluid （FF） levels of prostaglandin E2 （PGE2）, which plays a critical role within the ovary, and to investigate the effect of PGE2 on steroidogenesis in granulosa-lutein cells （GLCs） from women with and without endometriosis. Methods Thirty-three women with laparoscopically documented endometriosis and 40 controls undergoing in vitro fertilization （IVF） were studied. We assayed the concentrations of PGE2 in FF, the production of E2 and progesterone in FF and in culture medium, and the expression of steroidogenic acute regulatory protein （STAR） and CYP19A1 in GLCs with the intervention of PGE2. Results PGE2 and progesterone concentrations were increased and displayed positive correlation in endometriotic FE PGE2 induced the expression of STAR and the production of progesterone in GLCs from women with endometriosis, and the expression of STAR and the production of progesterone were increased in GLCs from women with endometriosis. However, there were no significant effects of PGE2 on promoting the production of E2 or the expression of CYP19A1 in GLCs. Moreover, the production of E2 and the expression of CYP19A1 in GLCs from women with endometriosis were significantly decreased compared to the controls. Conclusions PGE2 concentrations are increased in endometriotic FF, along with concomitant increases in progesterone and STAR. In contrast, the E2 and CYP19A1 are decreased in GLCs, which may delay the development of the follicles and cause an imbalance in the follicular steroid hormone levels. These changes may have close relationship with endometriosis-associated infertility.

How to interpret low human chorionic gonadotropin levels in cerebrospinal fluid of patients with intracranial germinoma？

Human chorionic gonadotropin （hCG） level in cerebrospinal fluid （CSF hCG） is very useful for thediagnosis of intracranial germinoma. We have previously reported on two patients with germinoma located in the basal ganglia. Their CSF hCG levels were extremely low upon presentation.1 However, significantly increased CSF hCG levels were detected when these two patients had a relapse. The experience acquired in these two cases is helpful to appropriately interpret the CSF hCG levels that are used to detect intracranial germinoma, evaluate the effectiveness of treatment and monitor recurrence.

Role of human visceral pleura in pleural fluid turnover:Need for morphological evidence of lymphatic stomata

To the Editor. We read with great interest the article by Gao and his colleagues proposing a new schema and hypothesis regarding pleural fluid turnover. In the proposed hypothesis, the visceral pleura possesses a more active role in humans than what was believed. The authors support the notion that in humans the pulmonary lymphatic vessels originate directly from the mesothelium of the visceral pleura and that it has an effect on both infiltration and reabsorption of pleural fluid in human beings.

B cell development and antibody responses in human immune system mice:current status and future perspective

Humanized immune system(HIS)mice have been developed and used as a small surrogate model to study human immune function under normal or disease conditions.Although variations are found between models,most HIS mice show robust human T cell responses.However,there has been unsuccessful in constructing HIS mice that produce high-affinity human antibodies,primarily due to defects in terminal B cell differentiation,antibody affinity maturation,and development of primary follicles and germinal centers.In this review,we elaborate on the current knowledge about and previous attempts to improve human B cell development in HIS mice,and propose a potential strategy for constructing HIS mice with improved humoral immunity by transplantation of human follicular dendritic cells(FDCs)to facilitate the development of secondary follicles.