Keys molecules implicated in apoptotic pathway of sea urchin embryos

Molecular Alterations in Spermatozoa of a Family Case Living in the Land of Fires. A First Look at Possible Transgenerational Effects of Pollutants.

In our previous work, we reported alterations in protamines/histones ratio, in DNA binding of these proteins and their involvement in DNA oxidative damage in 84% of the young men living in the Land of Fires. In the present work, we extended our findings, evaluating any alterations in spermatozoa of a family case, a father and son, living in this area, to also give a first look at the possibility of transgenerational inherited effects of environmental contaminants on the molecular alterations of sperm nuclear basic proteins (SNBP), DNA and semen parameters. In the father and son, we found a diverse excess of copper and chromium in the semen, different alterations in SNBP content and low DNA …

Sea urchin embryos cadmium exposed as an experimental model system for studying the relationship between autophagy and apoptosis

HPV in semen: evaluation of the link among infection, sperm parameters and viral genome integration

Human papillomaviruses (HPVs) are agents of the most common sexually transmitted diseases. Both in men and in women, HPVs cause a variety of clinical symptoms ranging from warts to cancer. It has been shown that infection with oncogenic HPV is the major cause of development of cervical cancer in women and less frequently of penile cancers in men. HPV infects epithelial cells and can bind other cell types. Its presence in semen is widely known, but the effects on fertility and reproductive function are still controversial. Partners of women with high-grade squamous intraepithelial lesions were enrolled. From each subject seminal parameters were evaluated. HPV was detected and genotyped both …

Sea Urchin Mitochondrial Matrix Contains a 56-kDa Chaperonine-like Protein

Abstract Paracentrotus lividus mitochondrial matrix contains a constitutive hsp of 56-KDa which cross reacts with a serum anti-hsp-60 chaperonine from yeast mitochondria. The localization of hsps preexisting or newly synthesized in different subcellular fractions of gastrula embryos is also analyzed by two-dimensional electrophoresis.

Anti-inflammatory properties of an aldehydes-enriched fraction of grapefruit essential oil

Chronic inflammation is linked to the development of numerous diseases and is accompanied by increased cytokine secretion. Macrophages provide a first line of defense against pathogens that under inflammatory stimuli release pro-inflammatory cytokines. The essential oil (EO) fractions obtained from Citrus spp. rich in different compounds have gained the attention of both researchers and users during the last decades. In particular, grapefruit (Citrus paradisi) peel is rich in phenolics and flavonoids with several health benefits, including anti-inflammatory actions. Additionally, its EO consists of a large number of compounds such as monoterpenes, sesquiterpenes, alcohols, aldehydes, esters…

Morte cellulare programmata in oociti umani dopo ICSI

Correlation between the DNA fragmentation index (DFI) and sperm morphology of infertile patients

Abstract Purpose To evaluate the correlation between the DNA Fragmentation Index (DFI) and sperm morphology in patients undergoing ICSI, as a predictive parameter in reproductive outcomes. Methods A retrospective study was conducted on 125 infertile patients enrolled in a fertility clinic. Seminal characteristics were measured following the WHO guidelines (2010) for the examination of the seminal fluid. After collecting motile sperm population by pellet swim up, DFI was calculated and simultaneously associated with sperm morphology using in situ TUNEL assay and an image analyzer software in at least 250 spermatozoa for each patient. Results All subjects were divided into two groups accordin…

Implication of specific isoforms of p38 in termothollerance and in the response to different stresses in sea urchin embryo

Achievement of thermotolerance through hsps phosphorylation in sea urchin embryos.

TPA treatment of sea urchin embryos is able to induce thermotolerance. Evidence is provided that TPA treatment induces phosphorylation of a constitutive stress protein of 38 KDa.

Sea urchin as in vivo model for the assessment of Manganese effect in embryos development

Molecular analyses on HPV infections in semen

Introduction: Human Papillomaviruses (HPVs) are non-enveloped double stranded DNA viruses classified in different genera and several different genotypes. Depending on the genotypes, HPV infections can be asymptomatic or can cause from warts to malignant tumours. Some genotypes, such as HPV18 and HPV16, are considered high risk (HR) HPVs and they are the major cause of cervical cancer. HPV is highly tissue-tropic and infects epithelial cells, but it can also binds other cell types. It is known that HPV can be found in semen but its effects on spermatozoa and male reproductive system are not completely clarified. Materials and Methods: In order to study the HPV infection in semen, we develope…

Selection of the best three oocytes for intracytoplasmic sperm injection (ICSI) using apoptotic analysis of cumulus cells

Introduction: We studied the apoptosis rate of the cumulus cells of individual cumulus-oocyte complex (COC), to verify a relationship with clinical outcomes, in terms of pregnancy and implantation rates. Usually oocytes are selected using morphological criteria. We tried to verify if cumulus cell apoptotic rate could be used as a molecular criteria in selecting oocytes with higher implantation potentiality. Material and Methods: The study design consisted in two different trials: in the first we investigated apoptosis rate in cumulus cells of the three selected oocytes, according to Italian law, to be fertilized by intracytoplasmic sperm injection (ICSI). In a second trial, average apoptosi…

Environmental factors as possible causes of DNA fragmentation in human sperms

Literature data demonstrated that some environmental factors could have a key role in the remarkable and continuous decline of sperm quality observed in the last fifty years. Specifically, in the Taranto area, data about the detrimental effects of environmental pollution are alarming because of the high level of poisons released in the atmosphere. Pollution coming from the plant causes health and fertility risks, mainly due to the exposure to the dioxin. Our study analyzed sperm samples from three patients groups: i) workers of local steel factories; ii) Taranto residents; iii) Controls. Results demonstrate that patients from the “factory workers” group, constantly exposed to environmental …

Cadmium effects on sea urchin embryos

Lower apoptosis rate in human cumulus cells, after r-LH administration in ovarian stimulation for IVF procedure

A relationship between human sperm morphology and DNA fragmentation in infertile patients

Pollution exposure and high DNA fragmentation index in human sperms: a case-control study

Study question: Could pollution exposure play a key role on sperm quality, in terms of DNA fragmentation index (DFI)? Summary answer: Patients exposed to high level of pollution show an higher percentage of sperm DNA fragmentation index in contrast with patient from control group. What is known already: Taranto area is characterized by a number of steel factories and petrochemical industries. Data about the detrimental effects of environmental pollution are alarming. Pollution coming from the industrial plants causes health and fertility risks, mainly due to the exposure to several pollutant (PM10, heavy metals, etc). Toxic substances can affect DNA directly or indirectly, through oxidative…

Sperm DNA fragmentation: An early and reliable marker of air pollution.

Environmental factors could have a key role in the continuous and remarkable decline of sperm quality observed in the last decades. This study compared the seminal parameters and sperm DFI in men living in areas with different levels of air pollution. Results demonstrate that both steel plants workers and patients living in a high polluted area show a mean percentage of sperm DNA fragmentation above 30%, highlighting a clear sperm damage. In this work, two different techniques were used to measure sperm DNA damage in patients’ groups, finding in both cases a high sperm DFI in patients living in polluted areas. We candidate sperm DNA fragmentation as a valuable early marker of the presence…

Autophagy is related to apoptosis in sea urchin embryos exposed to cadmium

The sea urchin embryo is a suitable model system that offers an excellent opportunity to investigate different defense strategies activated in stress conditions. We previously showed that cadmium treatment provokes the accumulation of metal in dose-time dependent manner in embryonic cells and the activation of defense systems, such as the synthesis of HSPs and/or the initiation of apoptosis. Analyzing autophagy, by neutral red, acridine orange and LC3-detection, we demonstrated that Cd-exposed embryos adopt this process as an additional stratagem to safeguard the developmental program. We observed that embryos treated with subletal Cd concentration activate massive autophagic response after…

Apoptosis rate in human germinal cells: quality assay.

Apoptosis is a process that eliminates superfluous or genetically compromised cells. Cell death is one of the most important aspects of organization of the developing, as alteration in timing, level, or pattern of cell death can lead to developmental anomalies. In our studies, we investigated apoptosis in follicular somatic and germ cells of patients with infertility problems. We found that cell death occur in human germinal vesicle, in some mature oocytes and in the oocytes that failed fertilization after ICSI (Intracytoplasmic Sperm Injection), this failure was strictly correlated with apoptosis. Still, after the Italian law number 40 of 2004, it was not available oocytes to research, and…

Effects of work exposure to electromagnetic waves and organic compounds on sperm DNA fragmentation in patients undergoing ICSI

Apoptosi in oociti umani non fecondati dopo ICSI

Sperm DNA integrity after conventional freezing vs vitrification with and without cryoprotectors

DNA fragmentation index, pAKT and pERK1/2 in cumulus cells are related to oocyte competence in patients undergoing in vitro fertilization programme

SummaryActivated pERK1/2 and pAKT are key players in supporting cell survival and proliferation pathways. Translocation of pERK1/2 into the nucleus, where it interacts with transcription factors and DNA itself, is instrumental in exerting an anti-apoptotic effect. In this study, pAKT levels, pERK1/2 nuclear localization and DNA fragmentation index (DFI) in cumulus cells of single cumulus–oocyte complexes of patients undergoing in vitro fertilization programmes were evaluated and correlated with the clinical outcome of the related embryos. For a positive clinical outcome of blastocyst development, pERK1/2 nuclear localization and DFI value had a significant inverse relationship, whereas the …

Relationship between apoptosis and survival molecules in human cumulus cells as markers of oocyte competence

SummaryTo select from a single patient the best oocytes able to reach the blastocyst stage, we searched for valuable markers for oocytes competence. We evaluated the DNA fragmentation index (DFI) and the level of some survival molecules, such as AKT, pAKT and pERK1/2, in individual cumulus cell–oocyte complexes (COC). The study included normo-responder women. The average age of the patients was 34.3. DFI in cumulus cells was evaluated using the terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labelling (TUNEL) assayin situ. AKT, pAKT and pERK1/2 were measured by immunological assay and densitometric analysis of fluorescent signals using NIS-Elements BR 3.10 image software. Statisti…

Autophagy is required for sea urchin oogenesis and early development.

SummaryAutophagy is a major intracellular pathway for the degradation and recycling of cytosolic components. Emerging evidence has demonstrated its crucial role during the embryo development of invertebrates and vertebrates. We recently demonstrated a massive activation of autophagy in Paracentrotus lividus embryos under cadmium stress conditions, and the existence of a temporal relationship between induced autophagy and apoptosis. Although there have been numerous studies on the role of autophagy in the development of different organisms, information on the autophagic process during oogenesis or at the start of development in marine invertebrates is very limited. Here we report our recent …

Lower sperm DNA fragmentation after r-FSH administration in functional hypogonadotropic hypogonadism

Abstract PURPOSE: An observational clinical and molecular study was designed to evaluate the effects of the administration of recombinant human FSH on sperm DNA fragmentation in men with a non-classical form of hypogonadotropic hypogonadism and idiopathic oligoasthenoteratozoospermia. METHODS: In the study were included 53 men with a non-classical form of hypogonadotropic hypogonadism and idiopathic oligoasthenoteratozoospermia. In all patients, sperm DNA fragmentation index (DFI), assessed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate (dUTP) in situ DNA nick end-labelling (TUNEL) assay, was evaluated before starting the treatment with 150 IU of recombinant hum…

Apoptosis rate in cumulus cells can be considerate as an indicator for the selection of embryos to improve ongoing pregnancy and implantation rate

L'apoptosi nelle cellule del cumulo ooforo dopo somministrazione di r-LH.

AN INNOVATIVE STRATEGY IN SELECTING OOCYTES WITH HIGH IMPLANTATION POTENTIALITY FOR INTRACYTOPLASMIC SPERM INJECTION PROCEDURE

The aim of the research was to investigate the apoptosis rate of individual cumulus cell-oocyte complexes (COC), associated to the level of pAKT, to verify the difference between oocytes who produce embryos able to reach the blastocyst stage compared with embryos arrested during the in vitro culture. It was demonstrated that DNA fragmentation in cumulus cells was remarkably lower in patients who achieved a pregnancy after ICSI cycles, related to the quality of oocytes and embryos1,2. AKT pathway plays a critical role in the regulation of cell survival, and most growth factors activate this pathway3. The study focused on 53 patients, involved after informed consent. In this prospective and r…

APOPTOTIC ANALYSIS OF CUMULUS CELLS FOR THE SELECTION OF COMPETENT OOCYTES TO BE FERTILIZED BY INTRACYTOPLASMIC SPERM INJECTION (ICSI)

Oocyte quality is one of the main factors for the success of in vitro fertilization protocols. Apoptosis is known to affect oocyte quality and may impair subsequent embryonic development and implantation. The aim of this study was to investigate the apoptosis rate of single and pooled cumulus cells of cumulus cell–oocyte complexes (COCs), as markers of oocyte quality, prior to intracytoplasmatic sperm injection (ICSI).We investigated the apoptosis rate by TUNEL assay (DNA fragmentation) and caspase-3 immunoassay of single and pooled cumulus cells of COCs. The results showed that DNA fragmentation in cumulus cells was remarkably lower in patients who achieved a pregnancy than in those who di…

The Histone Deacetylase Inhibitor JAHA Down-Regulates pERK and Global DNA Methylation in MDA-MB231 Breast Cancer Cells

The histone deacetylase inhibitor N-1-(ferrocenyl)-N-8-hydroxyoctanediamide (JAHA) down-regulates extracellular-signal-regulated kinase (ERK) and its activated form in triple-negative MDA-MB231 breast cancer cells after 18 h and up to 30 h of treatment, and to a lesser extent AKT and phospho-AKT after 30 h and up to 48 h of treatment. Also, DNA methyltransferase 1 (DNMT1), 3b and, to a lesser extent, 3a, downstream ERK targets, were down-regulated already at 18 h with an increase up to 48 h of exposure. Methylation-sensitive restriction arbitrarily-primed (MeSAP) polymerase chain reaction (PCR) analysis confirmed the ability of JAHA to induce genome-wide DNA hypomethylation at 48 h of expos…

Esiste una correlazione tra apoptosi e morfologia negli spermatozoi umani di pazienti infertili?

HUMAN SPERM CELL AS EARLY BIOINDICATOR OF MALE HEALTH AND REPRODUCTION

Environmental factors could have a key role in the remarkable and continuous decline of sperm quality observed in the last fifty years. This study addressed the gap of knowledge on the effect of air pollutants on sperm DNA fragmentation, comparing the seminal parameters from men living in locations with different levels of air pollution. The detrimental effects of environmental pollution in the Taranto area are alarming: the high level of pollutants released from the steel plants in the atmosphere can cause health and fertility issues. Our study analyzed sperm samples from three groups of patients: i) workers of Taranto steel plants; ii) Taranto residents; iii) Palermo residents, assumed as…

Apoptosis in Human Cumulus Cells

Autophagy during development of sea urchin embryos

Autophagy is a highly regulated mechanism that enhances cell eukaryotic survival under various environmental and cellular stresses, by breakdown and recycling of macromolecules and organelles. Here we report that in Paracentrotus lividus embryos autophagic process occur, at a lesser extend during physiological development and at greater levels after a cadmium treatment. By Acridine Orange staining, we found that embryonic cells exposed to cadmium display green fluorescence in cytoplasm and nucleus, and show considerable red fluorescent dots in cytoplasm. This evidence suggests formation of acidic auto-phagolysosomal vacuoles. By Neutral Red vital staining, specific for lysosomes and cellula…

Predictive Role of the p16 Immunostaining Pattern in Atypical Cervical Biopsies with Less Common High Risk HPV Genotypes

P16 immunostaining is considered a useful surrogate of transcriptionally active high-risk (hr) HPV infection. Only strong and widespread “block-like” immunoreactivity is considered specific, whereas weak/focal p16 positive immunostaining is considered not specific, and follow-up and HPV molecular detection is not indicated. The aim of the study was to evaluate the presence of HPV DNA and Ki67 immunostaining in 40 cervical atypical biopsies (CALs) with mild and focal histological features suggestive of HPV infection—20 cases with weak/focal p16 positive immunoreactivity and 20 cases negative for p16 expression. In 16/20 weak/focal p16 positive CALs (80%), the INNO-LiPA HPV genotyping detecte…

Potential impact of a nonavalent anti HPV vaccine in Italian men with and without clinical manifestations

AbstractHuman papilloma virus infection (HPV) is the most common sexually transmitted disease. Little is known about male infection. Nonavalent vaccine against types 6/11/16/18/31/33/45/52/58 was approved and neutral gender immunization programs have been proposed. This study evaluates the potential impact of nonavalent vaccine compared to quadrivalent in male living in Sicily (Italy). 58.7% of samples were HPV positive and forty-four types of HPV were identified. A significant higher estimated coverage of nonavalent vaccine than quadrivalent was observed (64.3% vs. 45.8%), with absolute and relative additional impact of 20.1% and 47.2%, respectively. Low impact of the vaccine were calculat…

Lower apoptosis rate in human cumulus cells after administration of recombinant luteinizing hormone to women undergoing ovarian stimulation for in vitro fertilization procedures.

Objective: To investigate the effects of recombinant (r-) LH supplementation in “low responder” patients undergoing ovarian stimulation with r-FSH for an IVF program. The apoptosis rate in cumulus cells was used as an indicator of oocyte quality. Design: Comparison of the rate of DNA fragmentation and caspase-3 activity in cumulus cells in women stimulated with r-LH and r-FSH, versus patients treated with r-FSH alone (control). Setting: In vitro fertilization (IVF) laboratory. Patient(s): Forty patients undergoing assisted fertilization programs treated with a GnRH agonist, or r-FSH treatment begun on day 3 of the cycle (control). In the r-LH group, from day 8 of gonadotropin stimulation, 1…

The autophagic demand during oogenesis and early development of sea urchin

The autophagic pathway is an evolutionarily conserved homeostatic process, responsible for degradation and recycling of long-term proteins and cytoplasmic organelles in eukaryotic cells. This process constitutively occurs at basal levels and is involved in cell survival. Increased autophagy is induced by environmental cues, such as starvation and many stress agents, while excessive levels of autophagy can lead to autophagic Programmed Cell Death, with features that differ from those of the apoptotic process. We recently demonstrated massive activation of autophagy in P. lividus embryos, in cadmium stress conditions, and the existence of a temporal relationship between induced autophagy end …

Potential role of p53 protein as a novel biomarker of sperm quality, able to predict the success of ART techniques. EcoFoodFertility Project

Introduction: Protein p53 is well known as “The guardian of genome”; it changes its concentration in human spermatozoa DNAin relation to the damage of the latter. It has been suggested thatthe role of the p53 ancestral gene was to ensure the integrity ofthe genomic germline and the fidelity of the evelopment process.The aim of this study is to evaluate if different concentrations of p53 protein in human spermatozoa could influence embryo quality and pregnancy rate and possibly representing a potential predictive marker of sperm quality for successful fertilization .Methods: From July 2013 to June 2017 we have examinatedretrospectively 79 couples with 2-5 years of infertility history.Males had…

Apoptosis in human unfertilized oocytes after ICSI

Apoptosi associata alla morfologia negli spermatozoi umani di pazienti infertili

p38 MAPK activity during sea urchin embryo development (Paracentrotus lividus)

Apoptosis rate of human cumulus cells from single oocytes in infertile patients as criteria for the selection of oocytes to be treated by intracytoplasmic sperm injection (ICSI).

The Role of Autophagy and Apoptosis During Embryo Development

Programmed cell death (PCD) and cell survival are two sides of the same coin. Autopha‐ gy and apoptosis are crucial processes during embryo development of Invertebrates and Vertebrates organisms, as they are necessary for the formation of a new organism, start‐ ing from a fertilized egg. Fertilization triggers cell remodeling from each gamete to a toti‐ potent zygote. During embryogenesis, the cells undergo various processes, thus allowing the transformation of the embryo into an adult organism. In particular, cells require the appropriate tools to suddenly modify their morphology and protein content in order to respond to intrinsic and external stimuli. Autophagy and apoptosis are involved…

Apoptosis in sea urchin embryos.

Abstract It is demonstrated by DNA electrophoresis analysis, morphological observations and TdT in situ reaction, that Paracentrotus embryos if treated with TPA plus heat undergo an apoptotic reaction. Indication is also obtained that non treated embryos undergo spontaneous apoptosis at the early pluteus stage, expecially in the districts of arms and intestine. The possible meaning of this latter observation is discussed.

SELECTION OF THE BEST OOCYTES FOR INTRACYTOPLASMICSPERM INJECTION (ICSI) USING APOPTOTIC ANALYSIS OF CUMULUS CELLS

Introduction: We studied the apoptosis rate of the cumulus cells of individual cumulus-oocyte complex (COC), to verify a relationship with clinical outcomes, in terms of pregnancy and implantation rates. Usually oocytes are selected using morphological criteria. We tried to verify if cumulus cell apoptotic rate could be used as molecular criteria in selecting oocytes with higher implantation potentiality (1;2). Materials and Methods: The study design consisted in two different trials: in the first, we investigated apoptosis rate in cumulus cells of the three selected oocytes, to be fertilized by intracytoplasmic sperm injection (ICSI); in a second trial, average apoptosis rate of the cumulu…

FSH administration reduces significantly sperm apoptosis only in the case of high DFI value: a study in idiopathic dispermic patients

Introduction: In the last decades sperm DNA quality has been recognized as one of the most important markers of male reproductive potential (Lewis and Aitken, 2005; Ozmen, 2007; Tarozzi, 2007), in contrast to standard semen parameters as sperm density, motility and morphology, which do not act as powerful discriminators between fertile and infertile men. DNA damage in the male germ line is a major contributor to infertility, miscarriage and birth defects in the offspring. In animal models, it has been unequivocally demonstrated that the genetic integrity of the male germ line plays a major role in determining the normality of embryonic development. In humans, many studies showed that sperm …

Evaluation of DNA damage in human spermatozoa: a sperm quality assay.

Several studies have demonstrated the relationship between gonadotrophine serum levels and the apoptosis rate in germinal cells. In human, lower FSH serum levels are associated with reduced Sperm Standard Parameters in term of concentration, motility and morphology and higher sperm DNA fragmentation. Hypogonadotrope hypogonadism is a clinical condition associated with male infertility, characterized by low serum levels of gonadotrophins. In this pilot study we investigated the spermatozoa quality of 11 patients with development of hypogonadotrope hypogonadism urdergoing in vitro fertilization programs. Recombinant-FSH administered to improve the sperm parameters, apoptosis rate, aiming to i…

Apoptosis in human unfertilized oocytes after intracytoplasmic sperm injection

Objective To investigate the presence of programmed cell death in unfertilized oocytes after intracytoplasmic sperm injection (ICSI), assuming that previous apoptotic events could be correlated with the fertilization failure. Design Comparison of the rate of DNA fragmentation in human oocytes at different stages of maturation soon after pick-up (control) and in unfertilized oocytes after ICSI treatment. Setting In vitro fertilization (IVF) laboratory with extensive ICSI experience. Patient(s) Sixty-three patients undergoing assisted fertilization by ICSI. Intervention(s) Terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling (TUNEL) assay and anticaspase-3 cleaved…

Gene Expression and Apoptosis Levels in Cumulus Cells of Patients with Polymorphisms of FSHR and LHB Undergoing in Vitro Fertilization Program

Background/Aims: FSH receptor (FSHR) Ala307Thr and Asn680Ser and LHβ chain (LHB) Trp28Arg and Ile35Thr polymorphisms affect the response to pharmacological ovarian stimulation with r-FSH in women undergoing assisted reproductive treatment (ART). Here, we evaluated the expression level of selected genes involved in follicle maturation and the possible onset of apoptosis in cumulus cells of patients with single and double FSHR and LHB polymorphisms, as potential markers of oocyte competence. Methods: Cumulus cells from 36 stimulated patients were collected and SNP genotyping performed by PCR. Gene expression was evaluated through real-time PCR, and apoptosis estimated via TUNEL assay, and cle…

Apoptosis rate in human spermatozoa: a sperm quality assay.

Several studies have demonstrated the relationship between gonadotrophine serum levels and the apoptosis rate in germinal cells (Bosco et al 2005). In human, lower FSH serum levels are associated with reduced Sperm Standard Parameters in term of concentration, motility and morphology and higher sperm DNA fragmentation. Hypogonadotrope hypogonadism is a clinical condition associated with male infertility, characterized by low serum levels of gonadotrophins. In this pilot study we investigated the spermatozoa quality of 11 patients with development of hypogonadotrope hypogonadism urdergoing in vitro fertilization programs. Recombinant-FSH administered to improve the sperm parameters, apoptosi…

A new method to select the best three oocytes to be treated for intracytoplasmic sperm injection (ICSI)

Apoptosis rate of cumulus cells can be considered as an indicator for the selection of embryos to improve ongoing pregnancy and implantation rates.

Cumulus cells apoptosis rate an adjunct to morphology evaluation for the embryo selection on day 3 could be considered a new tool, compared with embryo selection by morphology alone, to select the embryos with higher implantation potential to increase the clinical outcomes after ICSI. Several studies have demonstrated a lower cumulus cell apoptotic rates in women who achieved pregnancy compared with women who did not become pregnant after ICSI. A prospective randomized observational study on 76 ICSI patients was performed before Ovum Pick-Up. Patients were randomized into either the control group (embryo selection by morphology only, A group: 48 patients) or the treatment group (morphology …

New molecular markers for the evaluation of gamete quality.

Purpose: Only 30 % of IVF cycles result in a pregnancy, so that multiple embryos need to be replaced, per treatment cycle, to increase pregnancy rates, resulting in a multiple gestation rate of 25 %. The use of new markers in the gamete selection, could reduce the number of the oocytes to be fertilized and embryos to be produced, but the tools to evidence the gamete competence remain unavailable and more studies are needed to identify bio-markers to select the best oocyte and sperm to produce embryos with higher implantation potentiality. Methods: To define oocyte competence, the apoptosis of the surrounding cumulus cells and the oxygen consumption rates for individual oocytes before fertil…

Contact dermatitis due to nickel allergy in patients suffering from non-celiac wheat sensitivity

Background: Non‐celiac wheat sensitivity (NCWS) is a new clinical entity in the world of gluten‐related diseases. Nickel, the most frequent cause of contact allergy, can be found in wheat and results in systemic nickel allergy syndrome and mimics irritable bowel syndrome (IBS). Objective: To evaluate the frequency of contact dermatitis due to nickel allergy in NCWS patients diagnosed by a double‐blind placebo‐controlled(DBPC)challenge,and to identify the characteristics of NCWS patients with nickel allergy. Methods: We performed a prospective study of 60 patients (54 females, 6 males; mean age 34.1 ± 8.1 years) diagnosed with NCWS from December 2014 to November 2016; 80 age‐ and sex‐matched…

r-lh administration in multiple follicular stimolation for ivf: effects of apoptosis in cumulus cells, oocytes quality and clinical outcome

The effects of manganese on embryos of the sea urchin, Paracentrotus lividus: stress or not?.

Il fallimento della fecondazione: un evento predeterminato?

P–074 Chromatin Maturity Index (CMI) in unfixed and live spermatozoa and Aniline Blue (AB) stained as an additional evaluation parameter in idiopathic male infertility

Abstract Study question To investigate whether idiopathic male infertility may be due to the presence of histones in motile spermatozoa using a modified AB staining protocol. Summary answer No correlation between CMI in live motile spermatozoa, DNA Fragmentation Index (DFI) and other conventional seminal parameters were found in male infertile patients. What is known already The AB stain discriminates between lysine-rich histones and arginine/cysteine-rich protamines. Transition from histones to protamines during spermatogenesis remodels chromatin packaging and abnormalities in the substitution of those proteins maybe interfere with seminal parameters and affect male infertility. The correl…

p38 MAPK activation is required for Paracentrotus lividus skeletogenesis

We investigated the p38 MAPK role during sea urchin, Paracentrotus lividus, development. We found that at the morula stage, before the onset of skeletogenesis, p38 MAPK shows a peak of activity, and we tested whether p38 MAPK activity has any effect on skeletogenesis. By immunohistochemistry on whole-mount embryos we show the preferential localization of the active p38 form both in the presumptive PMCs and bilateral spiculo- genesis centers in control embryos, and in the radialized supernumerary spiculogenesis centers induced by NiCl2 treatment. By using SB203580, a p38 MAPK specific inhibitor, we show that p38 activity is required both for the initial triradiate spicule rudiments formation…

Effect of ovarian stimulation with recombinant LH/ FSH, recombinant FSH and human menopausal gonadotrophin (hMG) in reducing apoptosis rate in cumulus cells of patients undergoing ICSI. a pilot study

HPV infection in semen: results from a new molecular approach

AbstractHuman papillomavirus (HPV) is the agent of the most common sexually transmitted diseases causing a variety of clinical manifestations ranging from warts to cancer. Oncogenic HPV infection is the major cause of cervical cancer and less frequently of penile cancers. Its presence in semen is widely known, but the effects on fertility are still controversial. We developed a new approach to evaluate virus localisation in the different semen components. We analysed also the specific genotype localisation and viral DNA quantity by qPCR. Results show that HPV DNA can be identified in every fraction of semen: spermatozoa, somatic cells and seminal plasma. Different samples can contain the HP…

Autophagy as a defense strategy against stress: focus on Paracentrotus lividus sea urchin embryos exposed to cadmium

Autophagy is used by organisms as a defense strategy to face environmental stress. This mechanism has been described as one of the most important intracellular pathways responsible for the degradation and recycling of proteins and organelles. It can act as a cell survival mechanism if the cellular damage is not too extensive or as a cell death mechanism if the damage/stress is irreversible; in the latter case, it can operate as an independent pathway or together with the apoptotic one. In this review, we discuss the autophagic process activated in several aquatic organisms exposed to different types of environmental stressors, focusing on the sea urchin embryo, a suitable system recently in…

Sperm DNA integrity after conventional freezing vs vitrification with and without cryoprotectors in oligoastenospermic patients

Manganese effects on sea urchin embryos.

A new strategy in selecting oocytes using cumulus cells analysis of specific molecules of the apoptotic pathway, according to the ability to reach blastocyst stage.

Study question: The aim of the research was to investigate the apoptosis rate of individual cumulus cell–oocyte complexes (COC), associated to the level of p-Akt, to verify the difference between oocytes who produce embryos able to reach the blastocyst stage compared with embryos arrested during the in vitro colture. Summary answer: It was demonstrated that DNA fragmentation in cumulus cells was remarkably lower in patients who achieved a pregnancy after ICSI cycles, related to the quality of oocytes and embryos. Akt pathway plays a critical role in the regulation of cell survival, and most growth factors activate this pathway. What is known already: Studies on oocyte maturation have shown …

Apoptosi indotta da cadmio in embrioni di riccio di mare

Induction of skeletal abnormalities and autophagy in Paracentrotus lividus sea urchin embryos exposed to gadolinium.

Abstract Gadolinium (Gd) concentration is constantly increasing in the aquatic environment, becoming an emergent environmental pollutant. We investigated the effects of Gd on Paracentrotus lividus sea urchin embryos, focusing on skeletogenesis and autophagy. We observed a delay of biomineral deposition at 24 hours post fertilization (hpf), and a strong impairment of skeleton growth at 48 hpf, frequently displayed by an asymmetrical pattern. Skeleton growth was found partially resumed in recovery experiments. The mesodermal cells designated to biomineralization were found correctly migrated at 24 hpf, but not at 48 hpf. Western blot analysis showed an increase of the LC3-II autophagic marker…

Cadmium induces apoptosis in P. lividus embryos

Sperm p53 concentration: a potential new biomarker for environmental pollution. Preliminary data. (EcoFoodFertility Project)

Relazione tra autofagia e apoptosi in embrioni di riccio di mare esposti a stress

L’embrione di riccio di mare Paracentrotus lividus è stato utilizzato come organismo modello della biologia dello sviluppo per molti anni ed è considerato il deuterostoma più primitivo, con scheletro calcificato, correlato a protocordati e vertebrati. Tale sistema offre un'eccellente opportunità per studiare le numerose strategie di difesa che gli embrioni mettono in atto contro diverse condizioni di stress. In precedenza, abbiamo riportato che l’esposizione di embrioni a dosi citotossiche di cadmio, provoca l'accumulo intracellulare del metallo e l'attivazione del sistema difensivo, in modo dose-tempo dipendente, attraverso la sintesi di specifiche HSPs e/o l’innesco di apoptosi. Mediante …

Apoptotic pathway in sea urchin embryos

In human cumulus cells, the apoptotic rate may be considered an indicator for the selection of embryos to improve ongoing pregnancy and implantation rates

In order to identify the embryos with higher implantation potential to increase the clinical outcomes after ICSI, the apoptotic rate in human cumulus cells for the embryo selection on day 3, as an adjunct to morphology evaluation, could be considered a new tool compared with embryo selection by morphology alone. Several studies have demonstrated a lower cumulus cell apoptotic rate in women who achieved pregnancy compared with women who did not become pregnant after ICSI. A prospective randomized observational study on 76 ICSI patients was performed before Ovum Pick-Up. Patients were randomized into either the control group (embryo selection by morphology only, A group: 48 patients) or the t…

Sea urchin embryos exposed to cadmium as an experimental model for studying the relationship between autophagy and apoptosis

The sea urchin embryo is a suitable model that offers an excellent opportunity to investigate different defence strategies activated in stress conditions. We previously showed that cadmium accumulates in a dose- and time-dependent manner into embryonic cells, activating different stress and defence mechanisms, including the synthesis of HSPs and the onset of apoptosis and/or autophagy. In this paper we investigated the functional relationship between autophagy and apoptosis, evaluating apoptosis signals in cadmium-exposed Paracentrotus lividus embryos with inhibited autophagy. We found that the inhibition of autophagy produced the concurrent reduction of apoptosis, suggesting that the two p…

Apoptosis rate in human spermatozoa is not related to nuclear vacuoles

P–178 Mural granulosa cells of the human follicles indirectly show death molecular signals not depending on different ovarian stimulation protocols

Abstract Study question Could the expression of the anti-apoptotic molecules AKT, p-Akt and ERK1/2 in Mural Granulosa Cells (MGC) be considered as marker of oocyte quality? Summary answer MGCs activate cell death pathways in analyzed follicles and it is not influenced by different stimulation protocols and it is not correlated to oocyte competence. What is known already It has been previously demonstrated that apoptosis rate of mural granulosa or cumulus cells (CC) were correlated with follicular oocyte number, age, embryo numbers in IVF/ICSI and also clinical pregnancy. Moreover, our previous data demonstrated that in selected patients, who received recombinant LH associated with recombina…

Apoptosi in larve di Paracentrotus lividus.

Exposure to cytotoxic concentrations of manganese and effects on development

Manganese is one of the most abundant metals present in nature in trace amounts, which is accumulated and utilized by all forms of life. It plays a multitude of biological roles, but exposure of cells/organisms to high levels causes toxicity. In the marine environment, increasing concentrations of bio-available ions often result from anthropogenic activities, and, consequently, manganese represents a new emergent factor in environmental contamination. To study cyto-toxicity of manganese on development, we used an excellent model system as sea urchin embryos Paracentrotus lividus. In this work, we show results from two-dimensional gel electrophoresis (2DE) analysis that highlighted qualitati…

Apoptosi in cellule del cumulo come possibile bio-marcatore molecolare di competenza ovocitaria: risultati di uno studio pilota condotto su pazienti sottoposte a diversi protocolli di stimolazione ovarica.

L'apoptosi come criterio di valutazione comparativa tra attività LH e hCG.