scholarly journals Role of Tetraspanin CD9 Molecule in Fertilization of Mammals

2015 ◽  
pp. 279-293 ◽  
Author(s):  
J. JANKOVIČOVÁ ◽  
M. SIMON ◽  
J. ANTALÍKOVÁ ◽  
P. CUPPEROVÁ ◽  
K. MICHALKOVÁ
Keyword(s):  
Acrosome Reaction ◽  
Vitro Fertilization ◽  
Fertilization Process ◽  
Sperm Binding ◽  
Sperm Penetration ◽  
Ambiguous Data ◽  
Precise Role ◽  
Candidate Molecule

Fertilization process is a very clever and unique process comprising some essential steps resulting in formation of zygote. Tetraspanin CD9 is considered to be a serious candidate molecule participating in these events. The importance of CD9 has been discussed in relation to acrosome reaction, sperm-binding, sperm-penetration, sperm-egg fusion and eventually, egg activation. The abundant expression of CD9 oocyte plasma membrane and the presence of CD9-containing vesicles in the perivitelline space of intact oocytes have been confirmed. Despite the fact that majority of authors analyzed CD9 expressed on oocytes, several studies considered the function of sperm CD9, too. To understand CD9 involvement, various conditions of in vitro fertilization (IVF) assays using polyclonal as well as monoclonal antibodies or knockout mice were carried out. However, ambiguous data have been obtained about the importance of CD9 in sperm-egg binding or fusion. Although the current findings did not prove any hypothesis, the indispensable role of CD9 in fertilization process was not excluded and the precise role of CD9 remains unexplained.

317 ANTI-HYALURONIDASE ACTION OF ELLAGIC ACID EFFECTIVELY PREVENTS POLYSPERMY THROUGH SUPPRESSION OF THE ACROSOME REACTION INDUCED BY THE SPERM - ZONA INTERACTION DURING PORCINE IN VITRO FERTILIZATION

2007 ◽  
Vol 19 (1) ◽  
pp. 274
Author(s):  
I. Tokeshi ◽  
H. Tatemoto ◽  
N. Muto ◽  
T. Yoshimoto ◽  
S. Nakamura ◽  
...  
Keyword(s):  
Ellagic Acid ◽  
Acrosome Reaction ◽  
Radical Scavenging Activity ◽  
Radical Scavenging ◽  
Fluorescein Isothiocyanate ◽  
Cumulus Cells ◽  
Vitro Fertilization ◽  
Sperm Binding ◽  
Sperm Penetration

We previously reported that the anti-hyaluronidase agents oligosaccharide and tannic acid (TA) were efficient probes for promoting the normal fertilization process in terms of an effective decrease in the incidence of polyspermy, not only in cumulus-enclosed but also in denuded oocytes in pigs. It was unclear, however, why the polyspermic penetration into the zona pellucida (ZP) was directly prevented by the anti-hyaluronidase action. The present study was conducted to examine the effects of 3 tannin relatives [TA, gallic acid (GA), and ellagic acid (EA)] on IVF parameters and the acrosome reaction induced by the sperm–ZP interaction. The anti-hyaluronidase and radical-scavenging activities of tannin relatives were measured by the colorimetric and the DPPH methods, respectively. Porcine cumulus–oocyte complexes (COCs) were cultured for 44 h in 0.1 mL of TCM-199 supplemented with 0.6 mM cysteine, 40 mU mL-1 of FSH, 20 mU mL-1 of LH, and 10% porcine follicular fluid. After in vitro maturation (IVM), the COCs were freed from their cumulus cells and inseminated by frozen-thawed ejaculated sperm in modified Tris-buffered medium (IVF medium) containing 0 (control) or 5 �g mL-1 of tannin relatives. After 2 h of co-incubation, the oocytes were gently pipetted to remove loosely bound sperm and stained with Hoechst 33342 to count the number of ZP-bound sperm, or stained with fluorescein isothiocyanate (FITC)-PNA, PI, and 422,6-diamidino-2-phenylindole to evaluate the acrosomal status. At 10 h post-insemination, IVF parameters were examined by lacmoid staining. The data were analyzed by ANOVA and the Tukey-Kramer test. None of the tannin relatives caused a protective proteolytic modification of the ZP matrix or a reduction of the acrosomal proteolytic activity or the number of ZP-bound sperm. There was no difference in the sperm penetration rate even in the presence of tannin relatives (73-82%). However, the incidence of polyspermy was remarkably prevented by TA (32%; 31/98) and EA (21%; 20/94) compared with the control (58%; 58/100; P < 0.05), resulting from their strong anti-hyaluronidase actions, whereas GA without the anti-hyaluronidase action had no effect on the prevention of polyspermy (51%; 43/84). The rate of acrosome reaction induced by the sperm–ZP interaction was decreased by TA (15%; 123/833) and EA (16%; 110/708) compared with the control (25%; 238/939; P < 0.05), and a similar result was found in sperm binding to the pretreated ZP with 500 U of hyaluronidase for 2 h (18%; 351/1959). Interestingly, when sperm were incubated in IVF medium with 10 �g mL-1 of progesterone for 0.5 h to induce a compulsory acrosome reaction instead of the ZP, EA never disturbed the acrosome reaction (23%; 98/424) as control (23%; 102/437), although this reaction was blocked by TA (13%; 57/427) and GA (13%; 50/375), which possessed higher levels of radical-scavenging activity than EA (P < 0.05). These results indicate that the anti-hyaluronidase action of TA and EA effectively prevented polyspermy during porcine IVF as a consequence of suppression of the acrosome reaction functionally induced by the sperm–ZP interaction requiring the hyaluronidase intervention.

scholarly journals Sperm binding to ZP2-coated beads improve the efficiency of porcine in vitro fertilisation

Reproduction ◽  
2020 ◽  
Vol 160 (5) ◽  
pp. 725-735
Author(s):  
Julieta Gabriela Hamze ◽  
María Jiménez-Movilla ◽  
Raquel Romar
Keyword(s):  
Acrosome Reaction ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
In Vitro Fertilisation ◽  
Sperm Binding ◽  
Fertilisation Rate ◽  
Gamete Recognition ◽  
Boar Spermatozoa

The role of specific zona pellucida (ZP) glycoproteins in gamete interaction has not yet been elucidated in many species. A recently developed 3D model based on magnetic sepharose beads (B) conjugated to recombinant ZP glycoproteins (BZP) and cumulus cells (CBZP) allows the study of isolated ZP proteins in gamete recognition studies. The objective of this work was to study the role of porcine ZP2, ZP3 and ZP4 proteins in sperm binding, cumulus cell adhesion and acrosome reaction triggering. ZP protein-bound beads were incubated with fresh ejaculated boar spermatozoa and isolated cumulus cells for 24 h. The number of sperm bound to the beads, the acrosomal shrouds (presence of acrosomal content) on the bead’s surface, and the acrosome integrity (by means of PNA-FITC lectin) in bound and unbound sperm were studied. Finally, in vitro matured porcine oocytes mixed with BZP2 were inseminated in vitro using fresh sperm and fertilisation results evaluated. Over 60% of beads had at least one sperm bound after 2 h of coincubation. ZP2-beads (BZP2) and cumulus-ZP2-bead complexes (CBZP2) reached the highest number of sperm per bead, whereas BZP3 and BZP4 models showed the highest number of unbound reacted sperm cells and acrosomal shrouds. Fertilisation efficiency and monospermy rate increased when oocytes were fertilised in the presence of BZP2. We, therefore, conclude that in pigs, it is mainly ZP2 that is involved in sperm-ZP binding whereas ZP3 and ZP4 induce acrosome reaction. Using magnetic sepharose ZP2-bound beads might be a valuable tool to improve the fertilisation rate in pigs.

The influence of sperm concentration, length of the gamete co-culture and the evolution of different sperm parameters on the in vitro fertilization of prepubertal goat oocytes

Zygote ◽  
2010 ◽  
Vol 18 (4) ◽  
pp. 345-355 ◽  
Author(s):  
M.J. Palomo ◽  
T. Mogas ◽  
D. Izquierdo ◽  
M.T. Paramio
Keyword(s):  
Acrosome Reaction ◽  
Penetration Rate ◽  
Sperm Concentration ◽  
Semen Parameters ◽  
Oxygen Species ◽  
Vitro Fertilization ◽  
Rate Of Penetration ◽  
Sperm Penetration ◽  
Kinetics Of

SummaryThe aims of the present study were: (1) to evaluate the influence of sperm concentration (ranging from 0.5 × 106 to 4 × 106 spermatozoa/ml) and length of the gamete co-incubation time (2, 4, 6, 8, 10, 12, 16, 20, 24 or 28 h) on in vitro fertilization (IVF), assessing the sperm penetration rate; (2) to investigate the kinetics of different semen parameters as motility, viability and acrosome status during the co-culture period; and (3) to analyse the effect of the presence of cumulus–oocytes complexes (COCs) on these parameters. To achieve these objectives, several experiments were carried out using in vitro matured oocytes from prepubertal goats. The main findings of this work are that: (1) in our conditions, the optimum sperm concentration is 4 × 106 sperm/ml, as this sperm:oocyte ratio (approximately 28,000) allowed us to obtain the highest penetration rate, without increasing polyspermy incidence; (2) the highest percentage of viable acrosome-reacted spermatozoa is observed between 8–12 h of gamete co-culture, while the penetration rate is maximum at 12 h of co-incubation; and (3) the presence of COCs seems to favour the acrosome reaction of free spermatozoa on IVF medium, but not significantly. In conclusion, we suggest that a gamete co-incubation for 12–14 h, with a concentration of 4 × 106 sperm/ml, would be sufficient to obtain the highest rate of penetration, reducing the exposure of oocytes to high levels of reactive oxygen species produced by spermatozoa, especially when a high sperm concentration is used to increase the caprine IVF outcome.

294 THE EFFECT OF PLASMIN ON THE IN VITRO FERTILIZATION ABILITY OF PORCINE GAMETES

2006 ◽  
Vol 18 (2) ◽  
pp. 254
Author(s):  
H.-H. Rhee ◽  
S.-J. Sa ◽  
H.-T. Cheong ◽  
B.-K. Yang ◽  
C.-K. Park
Keyword(s):  
In Vitro Fertilization ◽  
Zona Pellucida ◽  
Acrosome Reaction ◽  
Proteolytic Enzymes ◽  
Extracellular Proteins ◽  
Control Group ◽  
Sperm Viability ◽  
Vitro Fertilization ◽  
Sperm Binding

Plasminogen activators (PAs) are specific proteolytic enzymes that convert the inactive proenzyme plasminogen to plasmin. The plasmin formed is a nonspecific, potent protease that cleaves blood fibrin clots and several other extracellular proteins. The purposes of the present study were (1) to assess the effect of plamin on sperm viability and acrosome reaction (AR), (2) to examine the effect of plasmin on zona pellucida (ZP) solubility and the binding of sperm to ZP, and (3) to evaluate the effect of plasmin on fertilization responses, including penetration and incidence of polyspermy during in vitro fertilization in the pig. Ejaculated semen was collected from three mature Duroc boars by artificial vagina. The same three boars were used for all experiments. The oocyte maturation medium used was North Carolina State University-23 (NCSU-23) medium supplemented with 10% (v/v) porcine follicular fluid (pFF), 0.6 mM cysteine, 10 IU/mL human chorionic gonadotropin (hCG; Sigma-Aldrich Corporation, St. Louis, MO, USA), and 10 IU/mL pregnant mare's serum gonadotropin (PMSG; Sigma). Porcine spermatozoa, which were washed in Dulbecco PBS (Sigma), were resuspended and incubated in fertilization medium (mTBM) containing 0, 0.1, 1.0, 10.0, or 100.0 ng/mL plasmin (Sigma). Data were analyzed by ANOVA and Duncan's multiple-range test using the Statistical Analysis System (SAS Institute, Inc., Cary, NC, USA). The present study suggests that sperm viability was not affected by plasmin treatment. Also, addition of plasmin in doses ranging between 0.1 and 100.0 ng/mL for 2, 4, or 6 h to washed boar spermatozoa resulted in enhancement of acrosome reaction (AR), compared with untreated cells. Concentrations of 0 and 0.1 ng/mL plasmin (83 � 15 and 95 � 18 sperm/oocyte, respectively) had no effect on sperm binding, whereas 1.0 (123 � 21 sperm/oocyte), 10.0 (124 � 16 sperm/oocyte), and 100 ng/mL (124 � 15 sperm/oocyte) plasmin increased (P < 0.05) sperm binding, compared with the control. The zona pellucida solubility (zona digestion time) was significantly (P < 0.05) lower in medium with 1.0 (123 � 24 s), 10.0 (99 � 15 s), or 100.0 ng/mL (95 � 19 s) plasmin, compared with control (176 � 27 s). When porcine oocytes and spermatozoa were co-incubated in various concentrations of plasmin for 6 h, the penetration rate was significantly (P < 0.05) higher in medium with 1.0 ng/mL plasmin (77.5 � 3.1%), compared with control. However, there were no significant differences in the polyspermic rates and mean numbers of sperm (MNS)/oocyte among the groups treated with plasmin and the control group. We found that addition of plasmin to fertilization medium increases the percentage of acrosome-reacted spermatozoa and the sperm-binding ability of the pig ZP. These results suggest that plasmin may play a role in events related to fertilization in the pig.

Differential influence of recombinant non-glycosylated and glycosylated glycodelin on human sperm function: comparative studies with hamster spermatozoa

2001 ◽  
Vol 13 (3) ◽  
pp. 111 ◽  
Author(s):  
B. Dutta ◽  
R. Ain ◽  
P. B. Seshagiri ◽  
A. A. Karande
Keyword(s):  
In Vitro Fertilization ◽  
Acrosome Reaction ◽  
Human Sperm ◽  
Fertilization Rate ◽  
Sperm Function ◽  
Vitro Fertilization ◽  
Placental Protein 14 ◽  
Placental Protein

Glycodelin, also known as placental protein 14, has been implicated in endometriosis-related infertility. To determine the role of glycodelin and its glycosylated state, the influence of recombinant nonglycosylated-glycodelin (nongly-glycodelin) and glycosylated-glycodelin (gly-glycodelin) on human sperm function was evaluated. Whereas there was a significant (P<0.001) increase in the capacitation of nongly-glycodelin-treated spermatozoa compared with untreated controls (28.8 1.0% v. 21 1.5% respectively), treatment of spermatozoa with gly-glycodelin markedly (P<0.001) inhibited capacitation (10.7 0. 3%); acrosome reaction (AR) remained unaltered in all treatments. In a zona-free hamster egg penetration assay, the egg penetration index was higher (P<0.001) with nongly-glycodelin-treated spermatozoa (3.4 0.3) than with gly-glycodelin-treated spermatozoa (0.4 0.1) and untreated spermatozoa (1.6 0.2). A similar influence of glycodelin on capacitation was observed with hamster spermatozoa. However, the AR rate was higher (P<0. 01) in nongly-glycodelin-treated spermatozoa (39.4 1.6%) than in either gly-glycodelin-treated spermatozoa (19.3 2.0%) or untreated controls (30.0 1.2%). Moreover, the in vitro fertilization rate was significantly (P<0.01) higher with nongly-glycodelin treated-spermatozoa compared with untreated spermatozoa (77.5 2.3% v. 52.9 4.3%) and gly-glycodelin-treated spermatozoa (38.3 6.5%; P<0. 05). These results indicate that whereas nongly-glycodelin improves, gly-glycodelin inhibits, capacitation and fertilization potential of human and hamster spermatozoa, and that the glycosylation status of glycodelin determines its influence on sperm function.

Differences between antigenic determinants of pig and cat zona pellucida proteins

Reproduction ◽  
2000 ◽  
pp. 15-23 ◽  
Author(s):  
K Jewgenow ◽  
M Rohleder ◽  
I Wegner
Keyword(s):  
In Vitro Fertilization ◽  
Zona Pellucida ◽  
Antigenic Determinants ◽  
Vitro Fertilization ◽  
Contraceptive Vaccine ◽  
Sperm Binding ◽  
Dependent Inhibition ◽  
Dose Dependent Inhibition ◽  
Dose Dependent

Despite many efforts, the control of reproduction in feral cat populations is still a problem in urban regions around the world. Immunocontraception is a promising approach; thus the present study examined the suitability of the widely used pig zona pellucida proteins (pZP) for contraception in feral domestic cats. Purified zona pellucida proteins obtained from pig and cat ovaries were used to produce highly specific antisera in rabbits. Antibodies against pZP raised in rabbits or lions were not effective inhibitors of either in vitro sperm binding (cat spermatozoa to cat oocytes) or in vitro fertilization in cats, whereas antibodies against feline zona pellucida proteins (fZP) raised in rabbits showed a dose-dependent inhibition of in vitro fertilization. Immunoelectrophoresis, ELISA and immunohistology of ovaries confirmed these results, showing crossreactivity of anti-fZP sera to fZP and to a lesser extent to pZP, but no interaction of anti-pZP sera with fZP. It is concluded that cat and pig zonae pellucidae express a very small number of shared antigenic determinants, making the use of pZP vaccine in cats questionable. A contraceptive vaccine based on feline zona pellucida determinants will be a better choice for the control of reproduction in feral cats if immunogenity can be achieved.

Activation of bovine oocytes by protein synthesis inhibitors: new findings on the role of MPF/MAPKs†

2021 ◽  
Author(s):  
Cecilia Valencia ◽  
Felipe Alonso Pérez ◽  
Carola Matus ◽  
Ricardo Felmer ◽  
María Elena Arias
Keyword(s):  
Protein Synthesis ◽  
Kinase Activity ◽  
Cyclin B1 ◽  
Protein Synthesis Inhibitors ◽  
Vitro Fertilization ◽  
New Findings ◽  
Bovine Oocytes ◽  
Dependent Pathway

Abstract The present study evaluated the mechanism by which protein synthesis inhibitors activate bovine oocytes. The aim was to analyze the dynamics of MPF and MAPKs. MII oocytes were activated with ionomycin (Io), ionomycin+anisomycin (ANY) and ionomycin+cycloheximide (CHX) and by in vitro fertilization (IVF). The expression of cyclin B1, p-CDK1, p-ERK1/2, p-JNK, and p-P38 were evaluated by immunodetection and the kinase activity of ERK1/2 was measured by enzyme assay. Evaluations at 1, 4, and 15 hours postactivation (hpa) showed that the expression of cyclin B1 was not modified by the treatments. ANY inactivated MPF by p-CDK1Thr14-Tyr15 at 4 hpa (P &lt; 0.05), CHX increased pre-MPF (p-CDK1Thr161 and p-CDK1Thr14-Tyr15) at 1 hpa and IVF increased p-CDK1Thr14-Tyr15 at 17 hours postfertilization (hpf) (P &lt; 0.05). ANY and CHX reduced the levels of p-ERK1/2 at 4 hpa (P &lt; 0.05) and its activity at 4 and 1 hpa, respectively (P &lt; 0.05). Meanwhile, IVF increased p-ERK1/2 at 6 hpf (P &lt; 0.05); however, its kinase activity decreased at 6 hpf (P &lt; 0.05). p-JNK in ANY, CHX, and IVF oocytes decreased at 4 hpa (P &lt; 0.05). p-P38 was only observed at 1 hpa, with no differences between treatments. In conclusion, activation of bovine oocytes by ANY, CHX, and IVF inactivates MPF by CDK1-dependent specific phosphorylation without cyclin B1 degradation. ANY or CHX promoted this inactivation, which seemed to be more delayed in the physiological activation (IVF). Both inhibitors modulated MPF activity via an ERK1/2-independent pathway, whereas IVF activated the bovine oocytes via an ERK1/2-dependent pathway. Finally, ANY does not activate the JNK and P38 kinase pathways.

542 Objective Sleep Duration, Sleep Timing and Completion of In Vitro Fertilization Cycles; a Prospective Cohort Study

SLEEP ◽  
2021 ◽  
Vol 44 (Supplement_2) ◽  
pp. A214-A214
Author(s):  
Chawanont Pimolsri ◽  
Xiru Lyu ◽  
Cathy Goldstein ◽  
Chelsea Fortin ◽  
Sunni Mumford ◽  
...  
Keyword(s):  
Embryo Transfer ◽  
Sleep Duration ◽  
Medical Center ◽  
Working Women ◽  
Vitro Fertilization ◽  
Circadian Misalignment ◽  
Logistic Regression Models ◽  
Sleep Timing

Abstract Introduction Sleep duration and circadian misalignment have been linked to fertility and fecundability. However, sleep in women undergoing IVF has rarely been examined. This study investigated the role of sleep duration and timing with completion of an IVF cycle. Methods Prospective study of women undergoing IVF at a tertiary medical center between 2015 and 2017. Sleep was assessed by wrist-worn actigraphy 1–2 weeks prior to the initiation of their IVF cycle. Reproductive profile, IVF cycle details, demographic and health information were obtained from medical charts. Sleep duration, midpoint and bedtime were examined in relation to IVF cycle completion using logistic regression models, adjusted for age and anti-Müllerian hormone levels. A sub-analysis excluded women who worked non-day shifts to control for circadian misalignment. Results A total of 48 women were studied. Median age was 33y (range 25–42), with 29% of women older than 35 years. Ten women had an IVF cycle cancellation prior to embryo transfer. These women had shorter sleep duration, more nocturnal awakenings, lower sleep efficiency, and later sleep timing in comparison to those who completed their cycle. Twenty-minute increases in sleep duration were associated with lower odds of an uncompleted IVF cycle (OR = 0.88; 95% CI 0.78, 1.00). Women with later sleep midpoints and later bedtime had higher odds of an uncompleted cycle relative to those with earlier midpoints and earlier bedtime; OR=1.24; 95% CI 1.09, 1.40 and OR=1.33; 95% CI 1.17, 1.53 respectively, per 20-minute increments. These results were independent of age, levels of anti-Müllerian hormone, or sleep duration, and remained unchanged after exclusion of shift-working women. Conclusion This study demonstrated the influence of sleep duration and sleep timing on the odds of an uncompleted IVF cycle prior to embryo transfer. Sleep is a modifiable behavior that may contribute to IVF cycle success. Support (if any):

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