Leptin is involved in acrosome reaction by facilitating activation of MAPK cascades in the Chinese mitten crab, Eriocheir sinensis

2020 ◽  
Vol 70 (1) ◽  
pp. 81-95
Author(s):  
Qing Li ◽  
Haitao Zhao ◽  
Lin He ◽  
Hongdan Yang ◽  
Qun Wang
Keyword(s):  
Acrosome Reaction ◽  
Calcium Ionophore ◽  
Eriocheir Sinensis ◽  
Calcium Ionophore A23187 ◽  
Mitogen Activated Protein Kinase ◽  
Ionophore A23187 ◽  
Mapk Cascades ◽  
Mitten Crab ◽  
Mitogen Activated Protein

Abstract The role of leptin has been documented in several studies, including activated threonine phosphorylation of extracellular signal-regulated kinase (ERK1/2) in the reproduction of rodents and humans. Our previous studies have demonstrated that mitogen-activated protein kinase (MAPK) cascades ERK, P38, and c-Jun N-terminal kinase (JNK) are involved in the spermatogenesis and acrosome reaction of Eriocheir sinensis. Therefore, the aim of this study was to investigate the expression of leptin and its receptor (LepR), and the effect of leptin on MAPK cascades during calcium ionophore A23187-induced spermatozoa acrosome reaction in crabs. Successful western blotting revealed a 16 kDa band for leptin, and 120 kDa and 90 kDa bands for the obese receptor (LepR), respectively, in the tested male reproductive tissues. Both leptin and LepR were localized at the pro-acrosomal vesicle and apical cap (AC) of spermatids, suggesting their role in the subsequent acrosome reaction. Moreover, acrosome reaction can be enhanced by leptin, and this effect decreased due to the anti-LepR antibody. Afterwards, we investigated the effects of leptin on MAPK cascades. The results showed that leptin mainly activated the phosphorylation of ERK, P38 and JNK proteins in the apical cap during the acrosome reaction in crab spermatozoa. This study addresses the role of leptin on spermatozoa, and suggests that leptin may induce molecular changes associated with spermatozoa during acrosome reaction.

scholarly journals Mechanisms of Mitogen-Activated Protein Kinase Activation in Experimental Diabetes

1999 ◽  
Vol 10 (4) ◽  
pp. 738-745
Author(s):  
MIDORI AWAZU ◽  
KENJI ISHIKURA ◽  
MARIKO HIDA ◽  
MAKIKO HOSHIYA
Keyword(s):  
Protein Kinase ◽  
Mesangial Cells ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Mitogen Activated Protein Kinase ◽  
Ionophore A23187 ◽  
Mobility Shift ◽  
Early Diabetes ◽  
Mapk Activity ◽  
Mitogen Activated Protein

Abstract. Various growth factors and vasoactive substances are implicated in the pathogenesis of renal growth seen in early diabetes mellitus (DM). Mitogen-activated protein kinase (MAPK) is an important mediator of these extracellular stimuli. Protein kinase C (PKC), an enzyme known to be stimulated in DM, also activates MAPK. Thus, MAPK activity was examined in glomeruli from streptozotocin-induced DM rats. MAPK activity, measured as myelin basic protein kinase, was elevated by approximately 50% in DM versus controls (CON). Increased protein contents of p42mapk and p44mapk, as well as increased tyrosine phosphorylation and mobility shift of p42mapk, were also observed in DM. Tyrosine dephosphorylation of pp42mapk, on the other hand, assessed by incubating glomerular membrane with or without sodium orthovanadate (vanadate), was significantly diminished in DM. Protein expression of MAPK phosphatase-1 (MKP-1), a dual specificity phosphatase that inactivates MAPK, was approximately 60% of CON. Reduction in MKP-1 was reproduced in cultured mesangial cells grown under high glucose (30 mM; HG). The suppression of MKP-1 was PKC-dependent since incubation of HG cells with phorbol 12-myristate 13-acetate for 24 h abolished it. Furthermore, calcium ionophore A23187 reversed the suppression, suggesting that blunted Ca2+ signalling, characteristic of HG cells secondary to PKC stimulation, may be the cause. These results demonstrate that glomerular MAPK is activated in DM by multiple mechanisms i.e., increases in protein contents, increased phosphorylation, and decreased dephosphorylation of the enzyme due to suppression of MKP-1. These alterations may have an implication in the pathogenesis of diabetic nephropathy.

Acrosome stability in the spermatozoa of dasyurid marsupials

2008 ◽  
Vol 20 (2) ◽  
pp. 295 ◽  
Author(s):  
N. A. Czarny ◽  
K. E. Mate ◽  
J. C. Rodger
Keyword(s):  
Disulfide Bonds ◽  
Acrosome Reaction ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Macropus Eugenii ◽  
Sminthopsis Crassicaudata ◽  
Triton X ◽  
Physical And Chemical ◽  
Physical Challenge

The spermatozoa of most marsupials lack nuclear stabilising disulfide-bonded protamines found in eutherian mammals. However, disulfide stabilisation has been observed in the acrosome of macropodid (Macropus eugenii) and phalangerid (Trichosurus vulpecula) marsupials. As a result this organelle, which is normally fragile in eutherian mammals, is robust and able to withstand physical and chemical challenge in these marsupials. The present study examined acrosomal characteristics of the spermatozoa of three dasyurid marsupials; the fat-tailed dunnart (Sminthopsis crassicaudata), eastern quoll (Dasyurus viverrinus) and northern quoll (Dasyurus hallucatus). In all species examined Bryan’s staining demonstrated that significant acrosomal loss occurred following physical challenge with osmotic stress, cryopreservation without cryoprotectant and exposure to detergent (Triton-X). Bromobimane staining indicated that the acrosomes of dasyurids lacked stabilising disulfide bonds. As reported for the wallaby and possum, calcium ionophore (A23187) did not induce the acrosome reaction-like exocytosis in dasyurid spermatozoa but treatment with diacylglycerol (DiC8) caused significant acrosome loss at concentrations similar to those effective for other marsupials. The present study found that the spermatozoa of dasyurids are more sensitive to physical challenge than the previously-studied marsupials and we suggest that this is due to the absence of acrosomal stabilising disulfide bonds.

Tobacco smoke releases performed mediators from canine mast cells and modulates prostaglandin production

1992 ◽  
Vol 263 (1) ◽  
pp. L67-L72
Author(s):  
P. S. Thomas ◽  
R. E. Schreck ◽  
S. C. Lazarus
Keyword(s):  
Mast Cells ◽  
Tobacco Smoke ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Prostaglandin D2 ◽  
Ionophore A23187 ◽  
Temperature Dependent ◽  
Prostaglandin Production ◽  
Lungs And Airways

The role of an extract of tobacco smoke in activating mast cells was studied. With the use of isolated, canine mast cells as a model, we found that cigarette smoke solution (CSS) induced the release of the performed mediators histamine and tryptase from these cells in an energy- and temperature-dependent, non-cytotoxic manner. There was no requirement for extracellular calcium. Nicotine tartrate did not reproduce the effect of CSS. Interestingly, mast cells produced little prostaglandin D2 (PGD2) in response to the CSS, and there was a concentration-related inhibition of calcium ionophore A23187-induced PGD2 synthesis. This suggests at least two mechanisms acting on the mast cell: tobacco smoke can directly activate mast cells to release performed mediators and can simultaneously inhibit prostaglandin production. These observations suggest a mechanism by which mast cells may participate in the bronchospastic and proinflammatory changes seen in the lungs and airways of smokers.

Relocation of myosin and actin, kinesin and tubulin in the acrosome reaction of bovine spermatozoa

2009 ◽  
Vol 21 (2) ◽  
pp. 364 ◽  
Author(s):  
Ifigenia Oikonomopoulou ◽  
Hitesh Patel ◽  
Paul F. Watson ◽  
Peter D. Chantler
Keyword(s):  
Molecular Motors ◽  
Acrosome Reaction ◽  
Molecular Motor ◽  
Calcium Ionophore ◽  
Cell Types ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Sequence Of Events ◽  
Bovine Spermatozoa ◽  
Number Of Cells

The mammalian acrosome reaction is a specialised exocytotic event. Although molecular motors are known to be involved in exocytosis in many cell types, their potential involvement in the acrosome reaction has remained unknown. Here, it has been shown that actin is localised within the equatorial segment and in the marginal acrosomal ridge of the heads of unreacted bull spermatozoa. Myosins IIA and IIB are found within the anterior acrosomal margins of virtually all sperm cells and, less prominently, within the equatorial segment. Tubulin was detected in the equatorial segment and around the periphery of the acrosome while kinesin was prominent in the equatorial segment. After induction of the acrosome reaction by means of the calcium ionophore A23187, the number of cells exhibiting actin fluorescence intensity in the anterior acrosomal margin decreased four-fold and those displaying equatorial segment fluorescence decreased 3.5-fold; myosin IIA immunofluorescence decreased in intensity with most spermatozoa losing equatorial staining, whereas there was little change in the distribution or intensity of myosin IIB immunofluorescence, except for a ~20% decrease in the number of cells exhibiting acrosomal staining. Tubulin became largely undetectable within the head and kinesin staining spread rostrally over the main acrosome region. A possible sequence of events that ties in these observations of molecular motor involvement with the known participation of SNARE proteins is provided.

scholarly journals Nitric oxide and peroxynitrite trigger and enhance release of neutrophil extracellular traps

2019 ◽  
Vol 77 (15) ◽  
pp. 3059-3075 ◽  
Author(s):  
Aneta Manda-Handzlik ◽  
Weronika Bystrzycka ◽  
Adrianna Cieloch ◽  
Eliza Glodkowska-Mrowka ◽  
Ewa Jankowska-Steifer ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitrosative Stress ◽  
Calcium Ionophore ◽  
Neutrophil Extracellular Traps ◽  
Calcium Ionophore A23187 ◽  
Human Neutrophils ◽  
Ionophore A23187 ◽  
Inflammatory Reactions ◽  
Extracellular Traps

Abstract Despite great interest, the mechanism of neutrophil extracellular traps (NETs) release is not fully understood and some aspects of this process, e.g. the role of reactive nitrogen species (RNS), still remain unclear. Therefore, our aim was to investigate the mechanisms underlying RNS-induced formation of NETs and contribution of RNS to NETs release triggered by various physiological and synthetic stimuli. The involvement of RNS in NETs formation was studied in primary human neutrophils and differentiated human promyelocytic leukemia cells (HL-60 cells). RNS (peroxynitrite and nitric oxide) efficiently induced NETs release and potentiated NETs-inducing properties of platelet activating factor and lipopolysaccharide. RNS-induced NETs formation was independent of autophagy and histone citrullination, but dependent on the activity of phosphoinositide 3-kinases (PI3K) and myeloperoxidase, as well as selective degradation of histones H2A and H2B by neutrophil elastase. Additionally, NADPH oxidase activity was required to release NETs upon stimulation with NO, as shown in NADPH-deficient neutrophils isolated from patients with chronic granulomatous disease. The role of RNS was further supported by increased RNS synthesis upon stimulation of NETs release with phorbol 12-myristate 13-acetate and calcium ionophore A23187. Scavenging or inhibition of RNS formation diminished NETs release triggered by these stimuli while scavenging of peroxynitrite inhibited NO-induced NETs formation. Our data suggest that RNS may act as mediators and inducers of NETs release. These processes are PI3K-dependent and ROS-dependent. Since inflammatory reactions are often accompanied by nitrosative stress and NETs formation, our studies shed a new light on possible mechanisms engaged in various immune-mediated conditions.

In vitro induction of the acrosome reaction in ovine spermatozoa by calcium ionophore A23187

2003 ◽  
Vol 51 (1) ◽  
pp. 103-109 ◽  
Author(s):  
Ö. Uçar ◽  
T. J. Parkinson
Keyword(s):  
Phase Contrast ◽  
Acrosome Reaction ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Contrast Microscopy ◽  
The Relationship ◽  
In Vitro Induction ◽  
Artificial Vagina

The relationship between concentration of calcium ionophore A23187 and incubation time upon the proportion of spermatozoa undergoing acrosome reaction (AR) in vitro was investigated in rams from a commercial artificial insemination (AI) program. Two ejaculates were collected by artificial vagina from each of nine rams of three breeds (Finn Dorset, Charolais and Suffolk) aged 8-36months. Each ejaculate was diluted in a skimmed milk extender. Spermatozoa were thereafter incubated for 45 or 60min in modified Tyrode's medium (TALP) which contained either zero, 0.1 or 1.0µM/l A23187. After fixing in 10% formaldehyde, the number of spermatozoa that had undergone AR was determined by phase contrast microscopy. In pre-incubation samples, 21.3± 3.3% of spermatozoa had undergone AR. Percentages of acrosome reacted spermatozoa were significantly (P<0.001) increased after incubation with A23187. After incubation with 0.1µM/l A23187 for 45 and 60min there were 22.4±3.0% and 31.7±4.3% acrosome reacted spermatozoa, respectively. After incubation with 1.0µM/l A23187 for 45 and 60min there were 46.2±6.5% and 53.8±5.9% acrosome reacted spermatozoa, whilst corresponding numbers in control samples were 17.0±2.7% and 22.3±4.2%. There was also a significant (P<0.001) effect of individual animals upon the responses to different concentrations of A23187. These findings indicate that (i) A23187 can be used to assess the AR of ovine spermatozoa in vitro and (ii) there are effects of individual animals upon the proportion of spermatozoa undergoing AR.

Soybean trypsin inhibitor as a probe for the acrosome reaction in motile cynomolgus macaque sperm

Zygote ◽  
2000 ◽  
Vol 8 (2) ◽  
pp. 127-137 ◽  
Author(s):  
Theodore L. Tollner ◽  
Ashley I. Yudin ◽  
Gary N. Cherr ◽  
James W. Overstreet
Keyword(s):  
Trypsin Inhibitor ◽  
Acrosome Reaction ◽  
Cynomolgus Macaque ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Soybean Trypsin Inhibitor ◽  
Epifluorescence Microscopy ◽  
Ultrastructural Observation ◽  
Ionophore A23187 ◽  
Motile Sperm

Soybean trypsin inhibitor (SBTI) inhibits the catalytic activity of serine proteases, and has been shown to bind to acrosin, an acrosomal hydrolase which is not exposed on the surface of macaque sperm until after the acrosome reaction. Following activation with caffeine and dibutyryl cAMP, cynomolgus macaque sperm were induced to acrosome react with calcium ionophore A23187 in the presence of SBTI and were fixed for ultrastructural observation. Transmission electron microscopy (TEM) revealed secondary labelling of anti-SBTI-IgG with colloidal gold in association with the acrosomal matrix and fused membranes of sperm undergoing the acrosome reaction, but gold labelling was not observed on acrosome-intact sperm. When SBTI was conjugated with the fluorochrome Alexa 488, labelled (acrosome-reacted) sperm showed bright fluorescence that ranged from a patchy or punctate appearance to solid labelling over the region of the acrosomal cap. Following treatment with ionophore, the percentages of total acrosome-reacted sperm (motile and non-motile) as assessed with Alexa-SBTI, fluorescein isothiocyanate-conjugated Pisum sativum agglutinin (FITC-PSA), and TEM were 54.6%, 51.6% and 61.5%, respectively. Measures of acrosomal status with FITC-PSA and Alexa-SBTI were highly correlated (r = 0.94; n = 3). Macaque zonae pellucidae were co-incubated with activated sperm for 1 min and then rinsed in medium containing Alexa-SBTI and immediately observed with epifluorescence microscopy. The mean percentage of Alexa-SBTI-labelled (acrosome-reacted) motile sperm bound to the zona was 45.7 ± 14 (range: 22–80.4%; n = 4). Fewer than 1% of the motile sperm in suspension surrounding the zonae were acrosome-reacted. Alexa-SBTI had no effect on sperm motility, survival, or zona binding capability.

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