R. Pasquariello

Bio: R. Pasquariello is an academic researcher from University of Milan. The author has contributed to research in topics: Oocyte & SNP genotyping. The author has an hindex of 8, co-authored 26 publications receiving 204 citations. Previous affiliations of R. Pasquariello include Parco Tecnologico Padano & Colorado State University.

Design and validation of a 90K SNP genotyping assay for the water buffalo (Bubalus bubalis)

Abstract: Background The availability of the bovine genome sequence and SNP panels has improved various genomic analyses, from exploring genetic diversity to aiding genetic selection. However, few of the SNP on the bovine chips are polymorphic in buffalo, therefore a panel of single nucleotide DNA markers exclusive for buffalo was necessary for molecular genetic analyses and to develop genomic selection approaches for water buffalo. The creation of a 90K SNP panel for river buffalo and testing in a genome wide association study for milk production is described here. Methods The genomes of 73 buffaloes of 4 different breeds were sequenced and aligned against the bovine genome, which facilitated the identification of 22 million of sequence variants among the buffalo genomes. Based on frequencies of variants within and among buffalo breeds, and their distribution across the genome, inferred from the bovine genome sequence, 90,000 putative single nucleotide polymorphisms were selected to create an Axiom® Buffalo Genotyping Array 90K. Results This 90K “SNP-Chip” was tested in several river buffalo populations and found to have ∼70% high quality and polymorphic SNPs. Of the 90K SNPs about 24K were also found to be polymorphic in swamp buffalo. The SNP chip was used to investigate the structure of buffalo populations, and could distinguish buffalo from different farms. A Genome Wide Association Study identified genomic regions on 5 chromosomes putatively involved in milk production. Conclusion The 90K buffalo SNP chip described here is suitable for the analysis of the genomes of river buffalo breeds, and could be used for genetic diversity studies and potentially as a starting point for genome-assisted selection programmes. This SNP Chip could also be used to analyse swamp buffalo, but many loci are not informative and creation of a revised SNP set specific for swamp buffalo would be advised.

The Role of Resveratrol in Mammalian Reproduction

Abstract: Resveratrol is one of the most investigated natural polyphenolic compounds and is contained in more than 70 types of plants and in red wine. The widespread interest in this polyphenol derives from its antioxidant, anti-inflammatory and anti-aging properties. Several studies have established that resveratrol regulates animal reproduction. However, the mechanisms of action and the potential therapeutic effects are still unclear. This review aims to clarify the role of resveratrol in male and female reproductive functions, with a focus on animals of veterinary interest. In females, resveratrol has been considered as a phytoestrogen due to its capacity to modulate ovarian function and steroidogenesis via sirtuins, SIRT1 in particular. Resveratrol has also been used to enhance aged oocyte quality and as a gametes cryo-protectant with mainly antioxidant and anti-apoptotic effects. In males, resveratrol enhances testes function and spermatogenesis through activation of the AMPK pathway. Furthermore, resveratrol has been supplemented to semen extenders, improving the preservation of sperm quality. In conclusion, resveratrol has potentially beneficial effects for ameliorating ovarian and testes function.

Alterations in oocyte mitochondrial number and function are related to spindle defects and occur with maternal aging in mice and humans

Abstract: The objective of this work was to determine the role of mitochondria in the loss of oocyte quality with maternal aging. Our results show that mitochondrial DNA (mtDNA) copy number and function are reduced in eggs from aged mice after both in vivo and in vitro maturation. Higher incidences of spindle abnormalities were observed in old eggs. However, no correlation with egg ATP content was found. In vitro matured eggs from aged mice did not have a normal cortical distribution of active mitochondria and were subject to increased oxidative stress due to higher levels of reactive oxygen species and lower expression of glutamate-cysteine ligase, catalytic subunit (Gclc). Supplementation of antioxidants during in vitro maturation of old eggs mitigated this affect, resulting in increased mtDNA copy number and mitochondrial function, a mitochondria distribution pattern similar to young eggs, and improved chromosomal alignment. Eggs from women of advanced maternal age (AMA) had lower mitochondrial function than eggs from young women, although both age groups displayed a cortical distribution pattern of active mitochondria. In contrast to the mouse, human eggs from AMA women had higher mtDNA copy number than eggs from young women following in vitro maturation. In summary, oocytes of older females are more susceptible to perturbations in mitochondrial number and function, which are associated with increased spindle abnormalities and oxidative stress during in vitro maturation. These results demonstrate that oocyte mitochondria play a critical role in age-related infertility.

Apelin System in Mammary Gland of Sheep Reared in Semi-Natural Pastures of the Central Apennines.

Abstract: Sheep are the most bred species in the Central Italy Apennine using the natural pastures as a trophic resource and grazing activity is fundamental to maintain the grassland biodiversity: this goal can be reached only ensuring an economical sustainability to the farmers. This study aimed to investigate the apelin/apelin receptor system in ovine mammary gland and to evaluate the differences induced by food supplementation, in order to shed light on this system function. A flock of 15 Comisana x Appenninica adult dry ewes were free to graze from June until pasture maximum flowering (MxF). From this period to pasture maximum dryness (MxD), in addition to grazing, the experimental group (Exp) was supplemented with 600 g/day/head of cereals. Apelin and apelin receptor were assessed by Real-Time PCR and immunohistochemistry on the mammary glands of subjects pertaining to MxF, MxD and Exp groups. They were detected in alveolar and ductal epithelial cells. The pasture maximum flowering group showed significant differences in apelin expression compared with experimental and MxD groups. Apelin receptor expression significantly differed among the three groups. The reduced apelin receptor expression and immunoreactivity levels during parenchyma involution enables us to hypothesize that apelin receptor plays a modulating role in the system control.

A Detailed Study of Rainbow Trout (Onchorhynchus mykiss) Intestine Revealed That Digestive and Absorptive Functions Are Not Linearly Distributed along Its Length.

Abstract: To increase the sustainability of trout farming, the industry requires alternatives to fish-based meals that do not compromise animal health and growth performances. To develop new feeds, detailed knowledge of intestinal morphology and physiology is required. We performed histological, histochemical, immunohistochemical and morphometric analysis at typical time points of in vivo feeding trials (50, 150 and 500 g). Only minor changes occurred during growth whereas differences characterized two compartments, not linearly distributed along the intestine. The first included the pyloric caeca, the basal part of the complex folds and the villi of the distal intestine. This was characterized by a significantly smaller number of goblet cells with smaller mucus vacuoles, higher proliferation and higher apoptotic rate but a smaller extension of fully differentiated epithelial cells and by the presence of numerous pinocytotic vacuolization. The second compartment was formed by the proximal intestine and the apical part of the posterior intestine complex folds. Here we observed more abundant goblet cells with bigger vacuoles, low proliferation rate, few round apoptotic cells, a more extended area of fully differentiated cells and no pinocytotic vacuoles. Our results suggest that rainbow trout intestine is physiologically arranged to mingle digestive and absorptive functions along its length.

Impact of Oxidative Stress on Age-Associated Decline in Oocyte Developmental Competence.

Abstract: Reproductive capacity in women starts to decline beyond their mid-30s and pregnancies in older women result in higher rates of miscarriage with aneuploidy. Age-related decline in fertility is strongly attributed to ovarian aging, diminished ovarian reserves, and decreased developmental competence of oocytes. In this review, we discuss the underlying mechanisms of age-related decline in oocyte quality, focusing on oxidative stress (OS) in oocytes. The primary cause is the accumulation of spontaneous damage to the mitochondria arising from increased reactive oxygen species (ROS) in oocytes, generated by the mitochondria themselves during daily biological metabolism. Mitochondrial dysfunction reduces ATP synthesis and influences the meiotic spindle assembly responsible for chromosomal segregation. Moreover, reproductively aged oocytes produce a decline in the fidelity of the protective mechanisms against ROS, namely the ROS-scavenging metabolism, repair of ROS-damaged DNA, and the proteasome and autophagy system for ROS-damaged proteins. Accordingly, increased ROS and increased vulnerability of oocytes to ROS lead to spindle instability, chromosomal abnormalities, telomere shortening, and reduced developmental competence of aged oocytes.

Chromosome-level assembly of the water buffalo genome surpasses human and goat genomes in sequence contiguity

Abstract: Rapid innovation in sequencing technologies and improvement in assembly algorithms have enabled the creation of highly contiguous mammalian genomes. Here we report a chromosome-level assembly of the water buffalo (Bubalus bubalis) genome using single-molecule sequencing and chromatin conformation capture data. PacBio Sequel reads, with a mean length of 11.5 kb, helped to resolve repetitive elements and generate sequence contiguity. All five B. bubalis sub-metacentric chromosomes were correctly scaffolded with centromeres spanned. Although the index animal was partly inbred, 58% of the genome was haplotype-phased by FALCON-Unzip. This new reference genome improves the contig N50 of the previous short-read based buffalo assembly more than a thousand-fold and contains only 383 gaps. It surpasses the human and goat references in sequence contiguity and facilitates the annotation of hard to assemble gene clusters such as the major histocompatibility complex (MHC). Despite technological advances, chromosome-level assemblies of mammalian genomes are still rare. Here, the authors use PacBio, Chicago and Hi-C approaches to generate a highly contiguous and partially-phased genome assembly for the water buffalo, Bubalus bubalis

Impact of oxidative stress on male and female germ cells: implications for fertility

Abstract: Male and female germ lines are vulnerable to oxidative stress. In spermatozoa, such stress triggers a lipid peroxidation cascade that culminates in the generation of electrophilic lipid aldehydes that bind to DNA and a raft of proteins involved in the delivery of functionally competent cells. One set of targets for these aldehydes are the proteins of the mitochondrial electron transport chain. When this interaction occurs, mitochondrial ROS generation is enhanced leading to the sustained generation of oxidative damage in a self-perpetuating cycle. Such damage affects all aspects of sperm function including motility, sperm-egg recognition, acrosomal exocytosis and sperm-oocyte fusion. Oxidative stress in the male germ line also attacks the integrity of sperm DNA with potential impacts on the developmental capacity of embryos and the health and wellbeing of the offspring. Potential pathways of reactive oxygen species (ROS) generation in male germ cells could involve enhanced lipoxygenase activity, activation of NADPH oxidase and/or electron leakage from mitochondria. Similarly, in the female germ line, both the induction of oocyte senescence following ovulation and the deterioration of oocyte quality with maternal age appear to involve the generation of oxidative damage. In this case, the mitochondria appear to be a particularly important source of ROS compromising the viability and fertilizability of the oocyte and interfering with the normal segregation of chromosomes during meiosis. In light of these considerations, antioxidants should have some role to play in the preservation of reproductive function in both men and women; however, we still await appropriate trials to test this hypothesis.

The Role of Oxidative Stress and Natural Antioxidants in Ovarian Aging.

Abstract: The ovarian system comprises vital organs in females and is of great significance for the maintenance of reproductive potential and endocrine stability. Although complex pathogenesis undoubtedly contributes to ovarian aging, increasing attention is being paid to the extensive influence of oxidative stress. However, the role of oxidative stress in ovarian aging is yet to be fully elucidated. Exploring oxidative stress-related processes might be a promising strategy against ovarian aging. In this review, compelling evidence is shown that oxidative stress plays a role in the etiology of ovarian aging and promotes the development of other ovarian aging-related etiologies, including telomere shortening, mitochondrial dysfunction, apoptosis, and inflammation. In addition, some natural antioxidants such as quercetin, resveratrol, and curcumin have a protective role in the ovaries through multiple mechanisms. These findings raise the prospect of oxidative stress modulator-natural antioxidants as therapeutic interventions for delaying ovarian aging.