Detection of SPAM1 in the bovine oviductal fluid

2014 ◽  
Author(s):  
Acuna Omar Salvador ◽  
Maria Jimenez-Movilla ◽  
Ismael Vilella ◽  
Lourdes Jara ◽  
Sebastian Canovas ◽  
...  
Keyword(s):  
Oviductal Fluid

scholarly journals Deleted in malignant brain tumor 1 is secreted in the oviduct and involved in the mechanism of fertilization in equine and porcine species

Reproduction ◽  
2013 ◽  
Vol 146 (2) ◽  
pp. 119-133 ◽  
Author(s):  
Barbara Ambruosi ◽  
Gianluca Accogli ◽  
Cécile Douet ◽  
Sylvie Canepa ◽  
Géraldine Pascal ◽  
...  
Keyword(s):  
Brain Tumor ◽  
Western Blot ◽  
Immunohistochemical Analysis ◽  
Fertilization Rate ◽  
Malignant Brain Tumor ◽  
Estrus Cycle ◽  
Immunofluorescence Analysis ◽  
Oviductal Fluid ◽  
Porcine Spermatozoa

Oviductal environment affects preparation of gametes for fertilization, fertilization itself, and subsequent embryonic development. The aim of this study was to evaluate the effect of oviductal fluid and the possible involvement of deleted in malignant brain tumor 1 (DMBT1) on IVF in porcine and equine species that represent divergent IVF models. We first performed IVF after pre-incubation of oocytes with or without oviductal fluid supplemented or not with antibodies directed against DMBT1. We showed that oviductal fluid induces an increase in the monospermic fertilization rate and that this effect is canceled by the addition of antibodies, in both porcine and equine species. Moreover, pre-incubation of oocytes with recombinant DMBT1 induces an increase in the monospermic fertilization rate in the pig, confirming an involvement of DMBT1 in the fertilization process. The presence of DMBT1 in the oviduct at different stages of the estrus cycle was shown by western blot and confirmed by immunohistochemical analysis of ampulla and isthmus regions. The presence of DMBT1 in cumulus–oocyte complexes was shown by western blot analysis, and the localization of DMBT1 in the zona pellucida and cytoplasm of equine and porcine oocytes was observed using immunofluorescence analysis and confocal microscopy. Moreover, we showed an interaction between DMBT1 and porcine spermatozoa using surface plasmon resonance studies. Finally, a bioinformatic and phylogenetic analysis allowed us to identify the DMBT1 protein as well as a DMBT1-like protein in several mammals. Our results strongly suggest an important role of DMBT1 in the process of fertilization.

scholarly journals Replacement of Albumin by Preovulatory Oviductal Fluid in Swim-Up Sperm Preparation Method Modifies Boar Sperm Parameters and Improves In Vitro Penetration of Oocytes

Animals ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1202
Author(s):  
Sergio Navarro-Serna ◽  
Evelyne París-Oller ◽  
Ondrej Simonik ◽  
Raquel Romar ◽  
Joaquín Gadea
Keyword(s):  
Calcium Concentration ◽  
Preparation Method ◽  
Culture Media ◽  
Penetration Rate ◽  
Sperm Parameters ◽  
High Concentration ◽  
Sperm Preparation ◽  
Boar Sperm ◽  
Oviductal Fluid

More suitable and efficient methods to protect gametes from external harmful effects during in vitro handling can be achieved by adding preovulatory porcine oviductal fluid (pOF) to in vitro culture media. The objective of this study was to assess the swim-up procedure’s suitability as a sperm selection method using a medium supplemented with 1mg/mL BSA, 1% preovulatory pOF (v/v), 1% v/v pOF plus 1mg/mL BSA, and 5mg/mL BSA. After selection, various sperm parameters were studied, such as sperm recovery rate, sperm morphology, motility (by CASA), vitality, acrosome status and intracellular calcium (by flow cytometry) and ability to penetrate oocytes in vitro. Around 2% of sperm were recovered after swim-up, and the replacement of BSA by pOF showed a beneficial reduction of motility parameters calcium concentration, resulting in an increased penetration rate. The combination of albumin and oviductal fluid in the medium did not improve the sperm parameters results, whereas a high concentration of BSA increased sperm morphological abnormalities, motility, and acrosome damage, with a reduction of calcium concentration and penetration rate. In conclusion, the replacement of albumin by preovulatory oviductal fluid in the swim-up sperm preparation method modifies boar sperm parameters and improves the in vitro penetration of oocytes.

scholarly journals 325 INSULIN-LIKE GROWTH FACTOR-I CONCENTRATIONS IN OVIDUCT FLUID OF SUPEROVULATED EWES DURING THE PERI-OVULAR PERIOD

2005 ◽  
Vol 17 (2) ◽  
pp. 313
Author(s):  
M.A. Kakar ◽  
S. Maddocks ◽  
M.F. Lorimer ◽  
D.O. Kleemann ◽  
S.K. Walker
Keyword(s):  
Acetic Acid ◽  
Liquid Chromatography ◽  
Growth Factor ◽  
High Performance ◽  
Enzyme Linked Immunosorbent Assay ◽  
Research Station ◽  
Insulin Like Growth Factor ◽  
Igf I ◽  
Oviduct Fluid ◽  
Oviductal Fluid

This study examined the concentrations of insulin-like growth factor-1 (IGF-1) in oviduct fluid during the peri-ovular period as a reference for the establishment of optimal in vitro culture conditions for sheep embryos. Six mature ewes (4–5 years, 58–67 kg) of comparable body condition were fed a standard diet for two weeks before the start of fluid collection. Ewes were superovulated using conventional treatment involving a progestagen, FSH, and GnRH treatment. Oviducts were catheterized four days (which is sufficient time to recover from surgery) before collection of oviductal fluid, which started one day (Day 1) before the time of ovulation (Day 0) and continued until five days later (Day 5). Oviductal fluid was acidified by diluting into 0.8 M acetic acid/0.2 M trimethylamine, pH 2.8, mixed, and incubated to dissociate IGFs from IGF-binding proteins (IGFBPs). Following incubation, acidified fluid was centrifuged at 10,000g through a 0.1-mm Micro-spin centrifuge filter; the filtrate transferred to glass high-performance liquid chromatography (HPLC) vials. IGFs and IGFBPs were separated from one another by high-performance size-exclusion liquid chromatography using a Protein-Pak 125 column (Waters Corporation, Milford, MA, USA) and 0.2 M acetic acid, 0.05 M trimethylamine, pH 2.8, at a flow rate of 1 mL/min. Oviductal fluid IGF-I was collected in a single 2-mL fraction directly from the HPLC and its concentration measured by an IGF-I-specific enzyme-linked immunosorbent assay (Diagnostic Systems Laboratories, Inc. Webster, TX, USA). The data were analyzed by analysis of variance. The non-superovulation group had significantly higher concentrations of oviductal IGF-I compared with the superovulation group. In the superovulated group, there was, however, a significant effect of day on the oviductal fluid IGF-I concentration (P < 0.01) such that the concentrations of IGF-I first increased for three days and then decreased for the remaining four days. In the non-superovulation group, there was no significant two-way interaction between ovulation and day. It can be concluded that the levels of IGF-I increase over time and then decrease. Authors express thanks to the help of Jenn Skye and Hemish Turretfield Research Station SA.

scholarly journals Sperm Proteome after Interaction with Reproductive Fluids in Porcine: From the Ejaculation to the Fertilization Site

2020 ◽  
Vol 21 (17) ◽  
pp. 6060 ◽  
Author(s):  
Chiara Luongo ◽  
Leopoldo González-Brusi ◽  
Paula Cots-Rodríguez ◽  
Mª José Izquierdo-Rico ◽  
Manuel Avilés ◽  
...  
Keyword(s):  
Seminal Plasma ◽  
Proteome Analysis ◽  
Interacting Proteins ◽  
Sperm Transport ◽  
Two Fluids ◽  
Boar Sperm ◽  
Uterine Fluid ◽  
Proteome Changes ◽  
Oviductal Fluid

Ejaculated sperm are exposed to different environments before encountering the oocyte. However, how the sperm proteome changes during this transit remains unsolved. This study aimed to identify proteomic changes in boar sperm after incubation with male (seminal plasma, SP) and/or female (uterine fluid, UF; and oviductal fluid, OF) reproductive fluids. The following experimental groups were analyzed: (1) SP: sperm + 20% SP; (2) UF: sperm + 20% UF; (3) OF: sperm + 20% OF; (4) SP + UF: sperm + 20% SP + 20% UF; and (5) SP+OF: sperm + 20% SP + 20% OF. The proteome analysis, performed by HPLC-MS/MS, allowed the identification of 265 proteins. A total of 69 proteins were detected in the UF, SP, and SP + UF groups, and 102 proteins in the OF, SP, and SP + OF groups. Our results showed a higher number of proteins when sperm were incubated with only one fluid than when they were co-incubated with two fluids. Additionally, the number of sperm-interacting proteins from the UF group was lower than the OF group. In conclusion, the interaction of sperm with reproductive fluids alters its proteome. The description of sperm-interacting proteins in porcine species after co-incubation with male and/or female reproductive fluids may be useful to understand sperm transport, selection, capacitation, or fertilization phenomena.

scholarly journals Which Low-Abundance Proteins are Present in the Human Milieu of Gamete/Embryo Maternal Interaction?

2019 ◽  
Vol 20 (21) ◽  
pp. 5305 ◽  
Author(s):  
Canha-Gouveia ◽  
Paradela ◽  
Ramos-Fernández ◽  
Prieto-Sánchez ◽  
Sánchez-Ferrer ◽  
...  
Keyword(s):  
Embryo Development ◽  
Reproductive Tract ◽  
Culture Media ◽  
Adult Life ◽  
Early Embryo ◽  
Female Reproductive Tract ◽  
High Throughput Analysis ◽  
Maternal Interaction ◽  
Oviductal Fluid ◽  
Abundance Proteins

The improvement of the embryo culture media is of high relevance due to its influence on successful implantation rates, pregnancy, neonatal outcomes, and potential effects in adult life. The ideal conditions for embryo development are those naturally occurring in the female reproductive tract, i.e., the oviductal and uterine fluids. To shed light on the differences between chemical and natural media, we performed the first comparative study of the low abundance proteins in plasma, uterine, and oviductal fluid collected, simultaneously, from healthy and fertile women that underwent a salpingectomy. The rationale for this design derives from the fact that high-abundant proteins in these fluids are usually those coming from blood serum and frequently mask the detection of low abundant proteins with a potentially significant role in specific processes related to the embryo–maternal interaction. The proteomic analysis by 1D-nano LC ESI-MSMS detected several proteins in higher amounts in oviductal fluid when compared to uterine and plasma samples (RL3, GSTA1, EZRI, DPYSL3, GARS, HSP90A). Such oviductal fluid proteins could be a target to improve fertilization rates and early embryo development if used in the culture media. In conclusion, this study presents a high-throughput analysis of female reproductive tract fluids and contributes to the knowledge of oviductal and uterine secretome.

Heparin and dermatan sulphate induced capacitation of frozen-thawed bull spermatozoa measured by merocyanine-540

Zygote ◽  
2007 ◽  
Vol 15 (3) ◽  
pp. 225-232 ◽  
Author(s):  
A.-S. Bergqvist ◽  
J. Ballester ◽  
A. Johannisson ◽  
N. Lundeheim ◽  
H. Rodríguez-Martínez
Keyword(s):  
Chondroitin Sulphate ◽  
Heparan Sulphate ◽  
Negative Control ◽  
Dermatan Sulphate ◽  
Flow Cytometric ◽  
Merocyanine 540 ◽  
Incubation Duration ◽  
Flow Cytometric Study ◽  
Oviductal Fluid

SummaryGlycosaminoglycans (GAGs) are present in the oviduct in which the major part of sperm capacitation occurs. In this study we have tested how capacitation of frozen-thawed bull spermatozoa is effected by exposure to different GAGs detectable or possibly present in oviductal fluid; i.e. heparin, hyaluronan, heparan sulphate, dermatan sulphate and chondroitin sulphate. Following exposure of different duration, the spermatozoa were stained with either Chlortetracycline (CTC) or merocyanine-540 and evaluated with epifluorescent light microscopy or flow cytometry, respectively. Heparin elicited a significant increase in the number of alive, capacitated spermatozoa, either expressed as higher merocyanine-540 fluorescence (p < 0.0001) or as B-pattern (p = 0.0021) in the CTC assay, during 4 h of incubation. When comparing the different GAG treatments one by one to the negative control in the flow cytometric study, only heparin and dermatan sulphate were significant (p < 0.0001) higher than the control at 0–30 min of incubation. Duration of incubation did not affect the proportion of capacitated spermatozoa when measured as merocyanine-540 fluorescence or CTC B-pattern, but the length of the incubation did affect the number of dead (Yo-PRO 1 positive) spermatozoa (p < 0.0001). Exposure to zona pellucida proteins significantly increased the proportion of acrosome reacted spermatozoa (p = 0.016). Both heparin and dermatan sulphate induce capacitation of frozen-thawed bull spermatozoa in vitro.

Improvement of the developmental competence of canine oocyte using caffeine supplementation during IVM at different maturation time

Zygote ◽  
2018 ◽  
Vol 26 (2) ◽  
pp. 162-167 ◽  
Author(s):  
Mohamed Fathi ◽  
A. Salama ◽  
Magdy R. Badr
Keyword(s):  
Developmental Competence ◽  
Control Group ◽  
Free Medium ◽  
Maturation Time ◽  
Fluid Medium ◽  
Developmental Potential ◽  
Nuclear Maturation ◽  
Maturation Medium ◽  
Oviductal Fluid

SummaryThe aim of the current study was to investigate the effect of caffeine supplementation during in vitro maturation (IVM) for different maturation times on the developmental potential of canine oocytes recovered from ovariohysterectomized bitches. The recovered cumulus–oocytes complexes were in vitro matured for 72 h. Here, 10 mM caffeine was added to the maturation medium for different incubation times (caffeine from 0–72 h maturation, caffeine for the first 24 h of maturation only, caffeine addition from 24 to 48 h maturation time, caffeine addition from 48 to 72 h maturation or in caffeine-free medium, control group). The matured oocytes were in vitro fertilized using frozen–thawed spermatozoa. The presumptive zygotes were in vitro cultured in synthetic oviductal fluid medium for 5 days. The results showed that both maturation and fertilization rates were significantly higher (P ˂ 0.05) using caffeine-treated medium for the first 24 h of maturation compared with the control and other two groups of caffeine treatment (from 24 to 48 h and from 48 to 72 h), whereas use of caffeine-treated medium for a 0–72 h incubation time did not affect these rates (P > 0.05). Interestingly, the matured oocytes in caffeine-supplemented medium for the first 24 h or from 0–72 h showed a significant (P ˂ 0.05) increase in the total number of cleaved embryos compared with the control group. In conclusion, supplementation of the maturation medium with 10 mM caffeine for the first 24 h of maturation or during the whole maturation time (0–72 h) improved nuclear maturation and subsequent embryo development preimplantation following in vitro fertilization.

178 IMPROVED DEVELOPMENTAL COMPETENCE OF BOVINE EMBRYOS BY PGI2 ANALOG TREATMENT

2006 ◽  
Vol 18 (2) ◽  
pp. 197 ◽  
Author(s):  
B. S. Song ◽  
J. S. Kim ◽  
D. B. Koo ◽  
J. S. Park ◽  
K. K. Lee ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Culture Media ◽  
Cell Mass ◽  
Developmental Rate ◽  
Treated Group ◽  
Developmental Competence ◽  
Bovine Embryos ◽  
Frozen Semen ◽  
Oviductal Fluid

The microenvironment of the follopian tube, in which the oviductal fluid contains a variety of cytokines and growth factors, affects pre-implantation development of fertilized embryos in mammals. Prostaglandin I2 (PGI2, prostacyclin) exists in oviductal fluid and is synthesized from arachidonic acid by prostacyclin synthetase. PGI2 also enhances the implantation rate of mouse embryos. In this study, the effect of PGI2 analog on the development of bovine embryos was examined. Bovine cumulus oocytes complexes (COCs) were matured in TCM-199 medium supplemented with 10 IU/mL pregnant mare serum gonadotropin (PMSG), 10 IU/mL hCG, and 10 ng/mL epidermal growth factor (EGF) at 39�C, 5% CO2 in air for 20-22 h. Following in vitro maturation, COCs were fertilized in Fert-TALP medium containing 0.6% BSA using frozen semen. Also, oocytes matured in vitro were enucleated, individually reconstructed with bESF cells, fused, and then activated by treatment with 5 �M ionomycin for 5 min and 2 mM 6-DMAP for 4 h. In vitro-fertilized (IVF) and nuclear-transferred (NT) eggs were cultured in 50 ��L drops of CR1-aa medium supplemented with 0.3% BSA in the absence or presence of 1 �M PGI2 analog at 39�C, 5% CO2 in air, respectively. At 3 days of culture, cleaved embryos were further cultured in the same culture media supplemented with 10% FBS for 4 days. Allocations of blastocysts to inner cell mass (ICM) and trophoblast (TE) cells were investigated to assess embryo quality. All experiments were repeated more than three times. All data were analyzed by using the Duncan test of ANOVA by the Statistical Analysis System (SAS Institute, Inc., Cary, NC, USA) and numbers of nuclei in blastocysts were expressed as mean � SE. No difference was detected in the cleaved rate of the eggs between the treated- and nontreated groups. IVF zygotes treated with PGI2 analog represented a higher developmental rate (33%, 122/418) to the blastocyst stage than nontreated controls (24%, 107/456) (P < 0.05). Among IVF-derived blastocysts, interestingly, the proportion (46%, 84/181) of expanded blastocysts was significantly higher in the PGI2 analog-treated group compared with that in the nontreated group (28%, 46/164). The number of nuclei in (165 � 6.1, n = 15) in blastocysts in the PGI2 analog-treated group was higher than that (146.12 � 5.7, n = 18) in the nontreated group (P < 0.05). No difference was detected in the ratio of ICM to total cells between PGI2 analog-treated (42.0 � 3.0%) and nontreated groups (41.9 � 2.9%). Like the IVF embryos, NT embryos in the PGI2 analog-treated group showed a higher in vitro developmental rate (33.6%, 43/128) than the nontreated embryos (24.2%, 32/132) (P < 0.05). Our results indicate that PGI2 analog improves the kinetics of embryo development in cattle.

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