Capacitation of mouse spermatozoa. I. Correlation between the capacitation state and protein tyrosine phosphorylation

Development ◽  
1995 ◽  
Vol 121 (4) ◽  
pp. 1129-1137 ◽  
Author(s):  
P.E. Visconti ◽  
J.L. Bailey ◽  
G.D. Moore ◽  
D. Pan ◽  
P. Olds-Clarke ◽  
...  
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Tyrosine Phosphorylation ◽  
Bovine Serum ◽  
Female Reproductive Tract ◽  
Dependent Manner ◽  
Protein Tyrosine Phosphorylation ◽  
Sperm Capacitation ◽  
Protein Tyrosine

The molecular basis of mammalian sperm capacitation, defined functionally as those processes that confer on the sperm the acquisition of fertilization-competence either in vivo in the female reproductive tract or in vitro, is poorly understood. We demonstrate here that capacitation of caudal epididymal mouse sperm in vitro is accompanied by a time-dependent increase in the protein tyrosine phosphorylation of a subset of proteins of M(r) 40,000-120,000. Incubation of sperm in media devoid of bovine serum albumin, CaCl2 or NaHCO3, components which individually are required for capacitation, prevent the sperm from undergoing capacitation as assessed by the ability of the cells to acquire the pattern B chlortetracycline fluorescence, to undergo the zona pellucida-induced acrosome reaction and, in some cases, to fertilize metaphase II-arrested eggs in vitro. In each of these cases the protein tyrosine phosphorylation of the subset of capacitation-associated proteins does not occur. Protein tyrosine phosphorylation of these particular proteins, as well as sperm capacitation, can be recovered in media devoid of each of these three constituents (bovine serum albumin, CaCl2 or NaHCO3) by adding back the appropriate component in a concentration-dependent manner. The requirement of NaHCO3 for these phosphorylations is not due to an alkalinization of intracellular sperm pH or to an increase in media pH. Caput epididymal sperm, which lack the ability to undergo capacitation in vitro, do not display this capacitation-dependent subset of tyrosine phosphorylated proteins in complete media even after extended incubation periods, and do not fertilize metaphase II-arrested eggs in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)

Capacitation of mouse spermatozoa. II. Protein tyrosine phosphorylation and capacitation are regulated by a cAMP-dependent pathway

Development ◽  
1995 ◽  
Vol 121 (4) ◽  
pp. 1139-1150 ◽  
Author(s):  
P.E. Visconti ◽  
G.D. Moore ◽  
J.L. Bailey ◽  
P. Leclerc ◽  
S.A. Connors ◽  
...  
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Tyrosine Phosphorylation ◽  
Bovine Serum ◽  
Biologically Active ◽  
Dependent Manner ◽  
Protein Tyrosine Phosphorylation ◽  
Concentration Dependent Manner ◽  
Mouse Sperm ◽  
Protein Tyrosine

In the accompanying report (Visconti, P.E., Bailey, J.L., Moore, G.D., Pan, D., Olds-Clarke, P. and Kopf, G.S. (1995) Development, 121, 1129–1137) we demonstrated that the tyrosine phosphorylation of a subset of mouse sperm proteins of M(r) 40,000-120,000 was correlated with the capacitation state of the sperm. The mechanism by which protein tyrosine phosphorylation is regulated in sperm during this process is the subject of this report. Cauda epididymal sperm, when incubated in media devoid of NaHCO3, CaCl2 or bovine serum albumin do not display the capacitation-associated increases in protein tyrosine phosphorylation of this subset of proteins. This NaHCO3, CaCl2 or bovine serum albumin requirement for protein tyrosine phosphorylation can be completely overcome by the addition of biologically active, but not inactive, cAMP analogues. Addition of the active cAMP analogues to sperm incubated in media devoid of NaHCO3, CaCl2 or bovine serum albumin overcomes the inability of these media to support capacitation, as assessed by the ability of the cells to acquire the pattern B chlortetracycline fluorescence, to undergo the zona pellucida-induced acrosome reaction and, in some cases, to fertilize metaphase II-arrested eggs in vitro. The effects of the cAMP analogues to enhance protein tyrosine phosphorylation and to promote capacitation appears to be at the level of the cAMP-dependent protein kinase (PKA), since two specific inhibitors of this enzyme (H-89 and Rp-cAMPS) block the capacitation-dependent increases in protein tyrosine phosphorylation in sperm incubated in media supporting capacitation. Capacitation, as assessed by the aforementioned endpoints, also appears to be inhibited by H-89 in a concentration-dependent manner. These results provide further evidence for the interrelationship between protein tyrosine phosphorylation and the appearance of the capacitated state in mouse sperm. They also demonstrate that both protein tyrosine phosphorylation and capacitation appear to be regulated by cAMP/PKA. Up-regulation of protein tyrosine phosphorylation by cAMP/PKA in sperm is, to our knowledge, the first demonstration of such an interrelationship between tyrosine kinase/phosphatase and PKA signaling pathways.

scholarly journals Signal transduction mechanisms involved in in vitro ram sperm capacitation

Reproduction ◽  
2006 ◽  
Vol 132 (5) ◽  
pp. 721-732 ◽  
Author(s):  
Patricia Grasa ◽  
José Álvaro Cebrián-Pérez ◽  
Teresa Muiño-Blanco
Keyword(s):  
Tyrosine Phosphorylation ◽  
Calcium Influx ◽  
Calcium Entry ◽  
Calcium Entry Blocker ◽  
Protein Tyrosine Phosphorylation ◽  
Sperm Capacitation ◽  
Protein Tyrosine ◽  
Entry Blocker ◽  
Ram Sperm

We validate the chlortetracycline (CTC) technique for the evaluation of capacitation and acrosome reaction-like changes in ram sperm, carrying out a double estimation of the acrosome status after treatment with lysophosphatidylcholine, using fluorescein isocyanate (FITC)-RCA/ethidium homodimer 1 (EthD-1) and CTC/EthD-1. Highly consistent results and a positive correlation between the results of acrosome-reacted sperm evaluated with both techniques were obtained. In this study, we evaluate the effects of ram sperm capacitation of BSA, Ca2+, NaHCO3and cAMP agonists and their influence on the associated protein tyrosine phosphorylation. We found a time-dependent increase in capacitation related to protein tyrosine phosphorylation, either in the absence or the presence of BSA. The addition of an increasing concentration of cholesterol to samples containing BSA did not influence results. The effect of bicarbonate was concentration-dependent, with a significantly lowered value of non-capacitated sperm in the presence 18 and 25 mM. The addition of extracellular calcium did not significantly increase either the proportion of capacitated sperm or the protein tyrosine phosphorylation signalling, although a significantly higher value of acrosome-reacted sperm was found in samples containing 4 mM Ca2+. cAMP agonists increased capacitated sperm and protein tyrosine phosphorylation signalling. The inhibition of protein kinase A by H-89 caused a decrease in sperm capacitation. Addition of a calcium-entry blocker (Verapamil; Sigma) did not influence results, which suggests that the calcium entry blocker was unable to inhibit the calcium influx associated with capacitation in ram sperm. Our findings might benefit our understanding of the biochemical mechanisms involved in mammalian sperm capacitation and ultimately, fertility.

scholarly journals Evaluation of in vitro anti-inflammatory activity of the spadix of Colocasia affinis

2019 ◽  
Vol 19 (2) ◽  
pp. 30-34
Author(s):  
Vanlalhruaii ◽  
C. Malsawmtluangi ◽  
H. Lalhlenmawia
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum ◽  
Methanolic Extract ◽  
Inflammatory Activity ◽  
Dependent Manner ◽  
Protease Inhibition ◽  
Egg Albumin ◽  
Anti Inflammatory

The methanolic extract of C. affinis was evaluated for its in-vitro anti-inflammatory activity by bovine serum albumin denaturation method, egg albumin denaturation method and protease inhibition method at different concentrations. Diclofenac sodium was used as the reference drug. The extract exhibits anti-inflammatory activity in a concentration-dependent manner. In bovine serum albumin denaturation method, the extract at concentrations of 100, 200, 400, 600, 800, 1000 µg/ml showed 25.49, 28.43, 31.37, 37.25, 41.17, 48.03% inhibition respectively. In egg albumin denaturation method, the concentrations of 50, 100, 200, 300, 400, 500 µg/ml showed 5.3, 9.89, 28.12, 32.8, 43.2, 52.8% inhibition respectively. In protease inhibition method the extract at concentration of 100, 200, 300, 400, 500 µg/ml showed 8.87, 19.32, 28.56, 43.96, 59.92% inhibition respectively. Therefore, from the results it can be concluded that the methanolic extract of C. affinis possesses anti-inflammatory activity.

246 USE OF IONOPHORE A23187 ON BOVINE FROZEN SPERM INCREASES HYPERACTIVATED MOTILITY

2015 ◽  
Vol 27 (1) ◽  
pp. 212
Author(s):  
R. Garaguso ◽  
M. J. Franco ◽  
N. M. Ortega ◽  
T. Fanti ◽  
A. A. Mutto
Keyword(s):  
Tyrosine Phosphorylation ◽  
Female Reproductive Tract ◽  
Ionophore A23187 ◽  
Protein Tyrosine Phosphorylation ◽  
Sperm Capacitation ◽  
Control Groups ◽  
Protein Tyrosine ◽  
Motility Pattern ◽  
Phosphorylation Pattern ◽  
Bovine Spermatozoa

Sperm capacitation is critical for oocyte fertilization in mammals. The capacitated state is acquired by the spermatozoon during its passage through the female reproductive tract and can be induced in vitro. At a functional level, sperm capacitation is associated with changes in the motility pattern – the hyperactivated state (HA) – and terminates with the acrosome reaction (AR). These events are characteristically regulated by different Ca2+-signalling pathways. For this reason, Ca2+ ionophores, such as A23187, are commonly used in sperm capacitation studies. The induction of AR and IVF, as well as the assessment of protein phosporylation of tyrosine substrates are useful methods for the evaluation of the capacitation state of spermatozoa. Although the increase of protein tyrosine phosphorylation via PKA is associated with sperm capacitation, in the mouse, A23187 capacitates the spermatozoa, thus bypassing this pathway. The aim of the present work was to test the effect of the ionophore A23187 on acquisition of the capacitated state by evaluation of HA motility and protein phosphorylation pattern in frozen bovine spermatozoa. Motile bovine spermatozoa were isolated by gradient centrifugation (Percoll) from frozen/thawed samples and treated with different concentrations of A23187 (0.05, 0.1, 0.2, and 0.3 µM) in H-TYH medium without BSA. After 10 min of treatment, spermatozoa were washed in medium with BSA (11.2%) and incubated in H-TYH with BSA (6%) for 30 min. For control groups, sperm were incubated in H-TYH medium and DMSO. The motility pattern was visually identified and quantified using a computer-assisted sperm analysis system. The motile/immotile spermatozoa and the HA motility patterns of each group were statistically analysed by applying Fisher's tests. The protein tyrosine phosphorylation pattern was evaluated by a Western immunoblotting assay using heparin as a positive control of sperm capacitation. Our results showed that spermatozoa treated with A23187 had a significant increase in HA motility. The proportions of HA spermatozoa were 10.92, 31.27, and 75.4% for 0.1, 0.2, and 0.3 µM A23187, respectively (P < 0.05). On the other hand, the pattern of PKA-tyrosine phoshorylation characteristic of capacitated spermatozoa was absent after incubation with A23187, similar to the response seen in mouse spermatozoa. The percentage of motile spermatozoa in the control groups (H-TYH medium: 36% and DMSO: 27.95%; P < 0.05) was reduced as compared with that of the basal sperm suspension (65.4%, P < 0.05) after 30 to 40 min of incubation. Motility in the spermatozoa treated with 0.1 and 0.2 µM ionophore was similar – 65.9 and 68.1%, respectively, but it was reduced in the 0.3 group – 57.5% (P < 0.05). Thus, treatment with A23187 increased the percentage of spermatozoa with HA motility, probably suggesting an improvement in fertilizing capacity. Nevertheless, these promising results remain to be confirmed by IVF assay.

A Precipitin-like Reaction of the Hemolymph of the Lobster Homarus americanus

1969 ◽  
Vol 26 (5) ◽  
pp. 1392-1397 ◽  
Author(s):  
James E. Stewart ◽  
Diane M. Foley
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum ◽  
Serum Proteins ◽  
Homarus Americanus ◽  
Test Period ◽  
Fluorescent Material

The levels of fluorescent material in the hemolymph of lobsters injected with serum proteins from lobster hemolymph labelled with fluorescein remained relatively constant over a 6-day test period; the levels in lobsters injected with bovine serum albumin labelled with fluorescein declined rapidly. A precipitin-like reaction was observed when lobster hemolymph serum was titrated with bovine serum albumin in vitro.

Preparation and in vitro photodynamic activities of novel axially substituted silicon (IV) phthalocyanines and their bovine serum albumin conjugates

2006 ◽  
Vol 16 (9) ◽  
pp. 2450-2453 ◽  
Author(s):  
Xiong-Jie Jiang ◽  
Jian-Dong Huang ◽  
Yu-Jiao Zhu ◽  
Fen-Xiang Tang ◽  
Dennis K.P. Ng ◽  
...  
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum

Amuchina Gel Xgerm hand rub in vitro virucidal activity against SARS-CoV-2

2021 ◽  
Author(s):  
Alessandra Capezzone de Joannon ◽  
Angela Testa ◽  
Natalie Falsetto ◽  
Michela Procaccini ◽  
Lorella Ragni
Keyword(s):  
Detection Limit ◽  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum ◽  
Virucidal Activity ◽  
European Standard ◽  
Enveloped Viruses ◽  
Prevention Measure ◽  
Study Product

Aim: Ethanol is highly effective at inactivating enveloped viruses, including SARS-CoV-2. The aim of this study is to evaluate the virucidal activity of Amuchina Gel Xgerm (74% ethanol) against SARS-CoV-2, according to the European Standard EN14476:2013+A2:2019. Materials & methods: Virucidal activity of the study product was evaluated against SARS-CoV-2 strain USAWA1/2020 in suspension, in the presence of 0.3 g/l of bovine serum albumin. Results: The log10 reduction of SARS-CoV-2 in the presence of bovine serum albumin was ≥4.11 ± 0.12 after 30 s of exposure to the study product (80% dilution). Cytotoxicity was observed in the 100 dilution, affecting the detection limit by 1 log10. Conclusion: Virucidal activity against SARS-CoV-2 supports the effectiveness of this alcohol-based formulation as a prevention measure for COVID-19 illness.

scholarly journals In-vitro displacement interaction of atenolol and amlodipine on binding with bovine serum albumin when co-administered

2008 ◽  
Vol 2 (1) ◽  
Author(s):  
Md Ashraful Alam ◽  
Md Abdul Awal ◽  
Mahbub Mostofa ◽  
Md Kamrul Islam ◽  
Nusrat Subhan
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum

Bovine serum albumin (BSA) can replace patient serum as a protein source in an in vitro fertilization (IVF) program

1989 ◽  
Vol 6 (3) ◽  
pp. 164-167 ◽  
Author(s):  
Claudio A. Benadiva ◽  
Barbara Kuczynski-Brown ◽  
Tobi G. maguire ◽  
Luigi Mastroianni ◽  
George L. Flickinger
Keyword(s):  
In Vitro Fertilization ◽  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum ◽  
Protein Source ◽  
Vitro Fertilization

In Vitro Study of the Binding of Taxifolin to Bovine Serum Albumin and the Influence of Common Ions on the Binding

2010 ◽  
Vol 39 (4) ◽  
pp. 482-494 ◽  
Author(s):  
Xiaolei Shi ◽  
Xuwen Li ◽  
Yantao Sun ◽  
Wei Wei ◽  
Ruijie Yang ◽  
...  
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum ◽  
In Vitro Study ◽  
Vitro Study
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