4 SUBFERTILITY IN BULLS CARRYING A NONSENSE MUTATION IN TMEM95 IS DUE TO FAILURE TO PENETRATE THE ZONA PELLUCIDA

2017 ◽  
Vol 29 (1) ◽  
pp. 109 ◽  
Author(s):  
B. Fernandez-Fuertes ◽  
S. Kölle ◽  
P. Lonergan
Keyword(s):  
Reproductive Performance ◽  
Zona Pellucida ◽  
Nonsense Mutation ◽  
Genome Wide Association Study ◽  
Transmembrane Protein ◽  
Sperm Concentration ◽  
Sperm Function ◽  
Cleavage Rate ◽  
Cell Stage

In a recent genome-wide association study, male reproductive ability was assessed for 7962 Fleckvieh sires (Pausch et al. 2014 PLoS Genet. 10, e1004044). Forty bulls in the population with exceptionally poor reproductive performance were found to be homozygous for a nonsense mutation in the transmembrane protein 95 (TMEM95)-encoding gene. Ejaculates from these individuals exhibited normal sperm concentration, morphology, viability, and motility. However, out of 35,671 inseminations with these bulls, only 1.7% resulted in pregnancies. In addition, none of the bulls with normal reproductive performance was homozygous for this mutation. The aim of this study was to examine the effect of this nonsense mutation in TMEM95 on bovine sperm function in vitro. In all experiments, data were assessed for normality of distribution and analysed using a one-way ANOVA. The model included the main effects of treatment, replicate and their interactions. In Experiment 1, the fertilizing ability of sperm, assessed as oocyte cleavage rate, from 5 homozygous (mt/mt), 3 heterozygous (wt/mt), and 2 wildtype (wt/wt) Fleckvieh bulls was assessed. Oocytes fertilised with sperm from mt/mt males had a lower cleavage rate at 48 h post-fertilization than oocytes fertilised with sperm from wt/wt or wt/mt bulls (26 ± 3%, 72 ± 2%, and 79 ± 3%, respectively; P < 0.01). In addition, the kinetics of cleavage were slower in the mt/mt group as evidenced by a lower proportion of embryos beyond the 2-cell stage at 48 h (32 ± 12.6%, 90 ± 3.6%, and 87 ± 0.4%, respectively, P < 0.01). This translated into a lower blastocyst rate at Day 8 in the mt/mt group in comparison with wt/mt and wt/wt groups (2 ± 1%, 27 ± 3%, and 40 ± 2%, respectively, P < 0.01). In Experiment 2, fluorescent staining of oocytes 3 h after fertilization revealed a lower number of sperm from mt/mt bulls bound to the zona pellucida (ZP) in comparison with sperm from wt/wt or wt/mt individuals (3 ± 1.0, 11 ± 2.3, and 24 ± 9.4, respectively; P < 0.05). In order to further study the potential role of TMEM95 in sperm-ZP interaction, in Experiment 3 sperm from the 3 groups were used to fertilize ZP-free oocytes. Consistent with the previous results, sperm from mt/mt bulls were less able to penetrate oocytes with no ZP than sperm from wt/wt or wt/mt bulls (6 ± 2%, 44 ± 8%, and 68 ± 14%, respectively; P < 0.05). In conclusion, these results suggest that TMEM95 is involved in events that take place before ZP binding but are required for sperm to interact normally with the oocyte vestments and, ultimately, to achieve fertilization. Further study of the possible role of this protein in the initiation of sperm capacitation and hypermotility, as well as in early embryo development, will shed more light regarding the crucial role of TMEM95 in sperm function. Supported by the Irish Department of Agriculture, Food and The Marine under the Research Stimulus Programme (Grant No. 11 S 104).

The role of the zona pellucida in the development of mouse eggs in vivo

Development ◽  
1970 ◽  
Vol 23 (3) ◽  
pp. 539-547
Author(s):  
Jacek A. Modliński
Keyword(s):  
Zona Pellucida ◽  
Blastocyst Stage ◽  
Cell Stage ◽  
Development In Vitro ◽  
Single Blastomeres ◽  
Mammalian Eggs ◽  
Mouse Eggs

Up to the present time the function and significance of the zona pellucida in the development of mammalian eggs has not been fully explained. Zona-free mouse eggs will develop in vitro from the 2-cell stage, or later, up to the blastocyst stage (Tarkowski, 1961; Mintz, 1962; Gwatkin, 1963). Single blastomeres isolated at the 2-cell (Mulnard, 1965), 4- and 8-cell stage (Tarkowski & Wróblewska, 1967) will also develop in vitro up to the blastocyst stage. Similar experiments on development in vitro of 1- and 2-cell rabbit eggs (Edwards, 1964) showed that in this species also cleavage can occur when the zona pellucida is absent, although the blastomeres exhibit a tendency to fall away from each other. Tarkowski's observations (unpublished) would appear to show, however, that naked 1-, 2- and 4-cell mouse eggs do not develop when transferred to the oviduct. A few hours after transplanting the naked eggs none could be recovered by flushing the oviduct, whereas eggs surrounded by zonae which were transplanted simultaneously were recovered.

The role of insulin-like growth factor II and its receptor in mouse preimplantation development

2003 ◽  
Vol 15 (1) ◽  
pp. 37 ◽  
Author(s):  
M. Pantaleon ◽  
H. Jericho ◽  
G. Rabnott ◽  
P. L. Kaye
Keyword(s):  
Growth Factor ◽  
Growth Regulation ◽  
Negative Regulator ◽  
Insulin Like Growth Factor ◽  
Mouse Development ◽  
Cleavage Rate ◽  
Cell Stage ◽  
Factor Ii

Insulin-like growth factor II (IGF-II) and its receptor, the IGF-II/mannose-6-phosphate (IGF-II/M6P) receptor, are first expressed from the zygotic genome at the two-cell stage of mouse development. However, their role is not clearly defined. Insulin-like growth factor II is believed to mediate growth through the heterologous type 1 IGF and insulin receptors, whereas the IGF-II/M6P receptor is believed to act as a negative regulator of somatic growth by limiting the availability of excess levels of IGF-II. These studies demonstrate that IGF-II does have a role in growth regulation in the early embryo through the IGF-II/M6P receptor. Insulin-like growth factor II stimulated cleavage rate in two-cell embryos in vitro. Moreover, this receptor is required for the glycaemic response of two-cell embryos to IGF-II and for normal progression of early embryos to the blastocyst stage. Improved development of embryos in crowded culture supports the concept of an endogenous embryonic paracrine activity that enhances cell proliferation. These responses indicate that the IGF-II/M6P receptor is functional and likely to participate in such a regulatory circuit. The functional role of IGF-II and its receptor is discussed with reference to regulation of early development.

scholarly journals Prediction of Functional Consequences of Missense Mutations in ANO4 Gene

2021 ◽  
Vol 22 (5) ◽  
pp. 2732
Author(s):  
Nadine Reichhart ◽  
Vladimir M. Milenkovic ◽  
Christian H. Wetzel ◽  
Olaf Strauß
Keyword(s):  
Transmembrane Protein ◽  
Functional Characterization ◽  
Missense Mutations ◽  
Mutant Proteins ◽  
Functional Consequences ◽  
New Perspective ◽  
The Stability ◽  
Dynamics Simulations ◽  
And Function

The anoctamin (TMEM16) family of transmembrane protein consists of ten members in vertebrates, which act as Ca2+-dependent ion channels and/or Ca2+-dependent scramblases. ANO4 which is primarily expressed in the CNS and certain endocrine glands, has been associated with various neuronal disorders. Therefore, we focused our study on prioritizing missense mutations that are assumed to alter the structure and stability of ANO4 protein. We employed a wide array of evolution and structure based in silico prediction methods to identify potentially deleterious missense mutations in the ANO4 gene. Identified pathogenic mutations were then mapped to the modeled human ANO4 structure and the effects of missense mutations were studied on the atomic level using molecular dynamics simulations. Our data show that the G80A and A500T mutations significantly alter the stability of the mutant proteins, thus providing new perspective on the role of missense mutations in ANO4 gene. Results obtained in this study may help to identify disease associated mutations which affect ANO4 protein structure and function and might facilitate future functional characterization of ANO4.

A study of factors which influence the job satisfaction of stockpeople on commercial pig farms

1990 ◽  
Vol 1990 ◽  
pp. 95-95
Author(s):  
R C Segundo ◽  
P R English ◽  
G Burgess ◽  
S A Edwards ◽  
O MacPherson ◽  
...  
Keyword(s):  
Job Satisfaction ◽  
Reproductive Performance ◽  
Research Work ◽  
Farm Animals ◽  
Considerable Influence ◽  
Pig Farms ◽  
And Performance ◽  
The Uk ◽  
Farm Livestock

The role of stockmanship in relation to the wellbeing of farm animals has been emphasised in the UK Codes of Recommendations for the Welfare of Farm Livestock (MAFF, 1983). Moreover, previous research work has demonstrated important effects of good and bad stockmanship not only on welfare but also on growth, lactational and reproductive performance of pigs and other farm livestock (Hemsworth et al, 1987). There is a need, therefore, to establish the factors which motivate and demotivate stockpeople since the degree of job satisfaction is likely to have a considerable influence on the attitude and performance of stockpeople and on their empathy with the animals in their care. With this objective in mind, a questionnaire was designed to investigate the aspects which could have an influence on the job satisfaction of the stockpeople involved in pig production.

scholarly journals Calcium-free vitrification reduces cryoprotectant-induced zona pellucida hardening and increases fertilization rates in mouse oocytes

Reproduction ◽  
2006 ◽  
Vol 131 (1) ◽  
pp. 53-61 ◽  
Author(s):  
Mark G Larman ◽  
Courtney B Sheehan ◽  
David K Gardner
Keyword(s):  
Ethylene Glycol ◽  
Zona Pellucida ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Initial Increase ◽  
Cell Stage ◽  
Free Treatment ◽  
Zona Hardening ◽  
Free Media ◽  
Oocyte Freezing

Despite the success of embryo cyropreservation, routine oocyte freezing has proved elusive with only around 200 children born since the first reported birth in 1986. The reason for the poor efficiency is unclear, but evidence of zona pellucida hardening following oocyte freezing indicates that current protocols affect oocyte physiology. Here we report that two cryoprotectants commonly used in vitrification procedures, dimethyl sulfoxide (DMSO) and ethylene glycol, cause a large transient increase in intracellular calcium concentration in mouse metaphase II (MII) oocytes comparable to the initial increase triggered at fertilization. Removal of extracellular calcium from the medium failed to affect the response exacted by DMSO challenge, but significantly reduced the ethylene glycol-induced calcium increase. These results suggest that the source of the DMSO-induced calcium increase is solely from the internal calcium pool, as opposed to ethylene glycol that causes an influx of calcium across the plasma membrane from the external medium. By carrying out vitrification in calcium-free media, it was found that zona hardening is significantly reduced and subsequent fertilization and development to the two-cell stage significantly increased. Furthermore, such calcium-free treatment appears not to affect the embryo adversely, as shown by development rates to the blastocyst stage and cell number/allocation. Since zona hardening is one of the early activation events normally triggered by the sperm-induced calcium increases observed at fertilization, it is possible that other processes are negatively affected by the calcium rise caused by cryoprotectants used during oocyte freezing, which might explain the current poor efficiency of this technique.

The Role of the CX3CL1-CX3CR1 Axis in Renal Disease

2021 ◽  
Author(s):  
Diana Hamdan ◽  
Lisa A. Robinson
Keyword(s):  
Therapeutic Potential ◽  
Kidney Diseases ◽  
Transmembrane Protein ◽  
Soluble Form ◽  
Protective Effects ◽  
Chronic Kidney Diseases ◽  
The Family ◽  
Soluble Species ◽  
And Function

Excessive infiltration of immune cells into the kidney is a key feature of acute and chronic kidney diseases. The family of chemokines are key drivers of this process. CX3CL1 (fractalkine) is one of two unique chemokines synthesized as a transmembrane protein which undergoes proteolytic cleavage to generate a soluble species. Through interacting with its cognate receptor, CX3CR1, CX3CL1 was originally shown to act as a conventional chemoattractant in the soluble form, and as an adhesion molecule in the transmembrane form. Since then, other functions of CX3CL1 beyond leukocyte recruitment have been described, including cell survival, immunosurveillance, and cell-mediated cytotoxicity. This review summarizes diverse roles of CX3CL1 in kidney disease and potential uses as a therapeutic target and novel biomarker. As the CX3CL1-CX3CR1 axis has been shown to contribute to both detrimental and protective effects in various kidney diseases, a thorough understanding of how the expression and function of CX3CL1 are regulated is needed to unlock its therapeutic potential.

PIN-FORMED 1 regulates cell fate at the periphery of the shoot apical meristem

Development ◽  
2000 ◽  
Vol 127 (23) ◽  
pp. 5157-5165 ◽  
Author(s):  
T. Vernoux ◽  
J. Kronenberger ◽  
O. Grandjean ◽  
P. Laufs ◽  
J. Traas
Keyword(s):  
Cell Fate ◽  
Apical Meristem ◽  
Transmembrane Protein ◽  
Loss Of Function ◽  
Hybrid Identity ◽  
Hormone Transport ◽  
Early Markers ◽  
Ideal Tool ◽  
And Function

The process of organ positioning has been addressed, using the pin-formed 1 (pin1) mutant as a tool. PIN1 is a transmembrane protein involved in auxin transport in Arabidopsis. Loss of function severely affects organ initiation, and pin1 mutants are characterised by an inflorescence meristem that does not initiate any flowers, resulting in the formation of a naked inflorescence stem. This phenotype, combined with the proposed role of PIN1 in hormone transport, makes the mutant an ideal tool to study organ formation and phyllotaxis, and here we present a detailed analysis of the molecular modifications at the shoot apex caused by the mutation. We show that meristem structure and function are not severely affected in the mutant. Major alterations, however, are observed at the periphery of the pin1 meristem, where organ initiation should occur. Although two very early markers of organ initiation, LEAFY and AINTEGUMENTA, are expressed at the periphery of the mutant meristem, the cells are not recruited into distinct primordia. Instead a ring-like domain expressing those primordium specific genes is observed around the meristem. This ring-like domain also expresses a boundary marker, CUP-SHAPED COTYLEDON 2, involved in organ separation, showing that the zone at the meristem periphery has a hybrid identity. This implies that PIN1 is not only involved in organ outgrowth, but that it is also necessary for organ separation and positioning. A model is presented in which PIN1 and the local distribution of auxin control phyllotaxis.

P–430 Male fertility preservation: is there a role for cancer-induced inflammation that affects semen quality in oncological patients?

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
C Omes ◽  
V Tomasoni ◽  
R Bassani ◽  
V Amico ◽  
R E Nappi
Keyword(s):  
Oxidative Stress ◽  
Fertility Preservation ◽  
Semen Quality ◽  
Sperm Concentration ◽  
Semen Parameters ◽  
Cancer Type ◽  
Oncological Patients ◽  
Inflammatory State ◽  
Hematological Values

Abstract Study question What is the cause of semen quality impairment in oncological patients during fertility preservation programs? The cancer type and stadiation or the resulting inflammatory state? Summary answer The inflammatory state seems to be related to the decrease of sperm concentration, motility, morphology and viability due to the worsening of oxidative stress microenvironment. What is known already Fertility preservation acquired a great importance in the last decades due to increase survival of oncological patients, boost of diagnosis under 40 years and postponement of paternal age. At the time of cryopreservation, only one third of these males are normozoospermic. Tumor itself or other factors, added to psychological reasons, may be involved but there is no clear evidence. An imbalance of ROS (reactive oxygen species) in semen can compromise its quality. However, the correlation between cancer-related generalized stress state and fertility is poorly investigated. Inflammatory conditions induced by infections and pathologies, including cancer, increase ROS. Study design, size, duration Retrospective observational analysis was performed on 45 patients (29.0 ± 6.9 yrs) recruited during their fertility preservation program between 2016 and 2019 with written consent on use of their clinical data for research purpose. Patients presented several oncological diagnoses. Semen samples obtained from multiple collections (N = 58) were analyzed before applying standard freezing protocol. Data on semen parameters, inflammatory indices, hematological values and type/stage of tumors were collected. No exclusion criteria were applied. Participants/materials, setting, methods Routine semen analysis was performed according to the WHO standards. Sperm concentration and motility were evaluated on Makler Chamber, whereas eosin stain and Diff-quick slides were used for viability and morphology, respectively. Lymphoma was present in 72% of cases, leukemia in 8%, seminoma in 7% and other cancers in 13%. Correlations (Pearson/Spearman tests) among principal semen parameters and hematological values (leukocytes, erythrocytes, hemoglobin, RDW, albumin, etc.) were calculated with a P-value &lt;0.05 considered statistically significant. Main results and the role of chance The majority of semen samples showed a severe impairment, with one or more parameters under lower reference limits (WHO): 48.3% had sperm concentration under 15 millions/ml, 43.1% had a progressive motility under 32%, 41.4% had viability under 58% and 91.4% had abnormal morphology (under 4%). The role of potential inflammatory state was analyzed by correlating semen parameters and some hematological values. No correlation was found with cancer type. Negative association resulted between progressive motility (%PR) and leukocytes (p = 0.041) or RDW% (p = 0.015), but positive one with albumin (p = 0.012). Even sperm count, total motility (%PR+NP) and morphology were significantly correlated to RDW% (p = 0.003, p = 0.032, p = 0.034, respectively). These findings suggest a possible role of inflammation and ROS related generation in semen quality impairment. Indeed, albumin exerts a protective action, but leukocytes are known to cause ROS increase. Cancer-induced oxidative stress state may alter red blood cells homeostasis and vitality and increase erythrocytes turnover resulting in high RDW values. It is likely semen is worse when blood values indicate more severe cancer-induced inflammatory condition. Limitations, reasons for caution Significant correlations with type/stage of cancer were not found due to small number of each diagnosis, in spite our study considered 3 years of patients inclusion. Moreover, we lack to analyze the same patient before the cancer onset to avoid the influence of inflammatory state generated by the tumor itself. Wider implications of the findings: Understanding the influence of cancer-induced inflammatory state on semen quality could increase the awareness that clinicians should direct patient to the fertility preservation as soon as possible, even if diagnosis is still ongoing. It should be evaluated whether offering specific treatments may reduce oxidative stress conditions. Trial registration number Not applicable

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