Fertility Cryopreservation

The cryobiology is the science of low temperature biology. Fertility cryopreservation is a vital branch of reproductive science and involves the preservation of gametes (sperm and oocytes), embryos, and reproductive tissues (ovarian and testicular tissues) for use in assisted reproduction techniques (ART). The cryopreservation of reproductive cells is the process of freezing, storage, and thawing of spermatozoa or oocytes. It involves an initial exposure to cryoprotectants, cooling to subzero temperature, storage, thawing, and finally, dilution and removal of the cryoprotectants, when used, with a return to a physiological environment that will allow subsequent development. Proper management of the osmotic pressure to avoid damage due to intracellular ice formation is crucial for successful freezing and thawing procedure. Management of non-cryopreserved reproductive cells (i.e., spermatozoa or oocytes) and tissues (i.e., testicular tissue or ovarian tissue) is problematic due to difficulties in donor-recipient synchronization and the potential for transmission of infectious pathogens, which cumulatively limits widespread application of these techniques. Cryopreserved cells and tissues can endure storage for centuries with almost no change in functionality or genetic information, making this storage a method highly attractive. There is a pressing need for the development of optimum cryopreservation methods for reproductive cells and tissues from many species. There are two major techniques for cryopreservation: freeze-thaw processes and vitrification. The major difference between them is the total avoidance of ice formation in vitrification. However, the biotechnology of the reproduction, although widely implemented, has generated protocols currently used to cryopreserve bovine sperm or oocytes, for example, that are still suboptimal, and cannot readily be extrapolated to other species' gametes. ART provide an ensemble of strategies for preserving fertility in patients and commercially valuable or endangered species. Nevertheless, it is very difficult to successfully cryopreserve. Currently, there is a growing interest to understand the underlying cryobiological fundamentals responsible for low survival rates in an effort to develop better cryopreservation. The key factors that affect the life-span of spermatozoa are the combinations of storage temperature, cooling rate, chemical composition of the extender, cryoprotectant concentration, reactive oxygen species (ROS), seminal plasma composition and hygienic control. Sperm preservation protocols vary among animal species owing to their inherent particularities that change extenders used for refrigeration and freezing. On the other hand, oocytes are available only in limited number as compared to spermatozoa, therefore, a cryopreservation protocol must allow a high rate of viability maintenance when they are employed in practical application in ART programs. One of the key factors that influence the freezing process is the ratio of surface area to volume. The oocytes require a longer time to reach osmotic balance with the cryoprotectant solution than the spermatozoa, due to their bigger volume. Then, during cooling of oocytes, various forms of cellular damage may occur, including cytoskeleton disorganization, chromosome and DNA abnormalities, spindle disintegration, plasma membrane disruption and premature cortical granule exocytosis with its related hardening of the zona pellucida. Therefore, animal gametes have been shown to survive storage at low temperatures, and recent results are very encouraging, although reproducible methods have yet to be obtained in many species

Influence of ROS on Ovarian Functions

High level of ROS (Reactive Oxygen Species), due to an increased production of oxidant species and/or a decreased efficacy of antioxidant system, can lead to oxidative stress (OS) an emerging health risk factor involved in the aging and in many diseases, either in humans or in animals. ROS are a double-edged sword – they serve as key signal molecules in physiological processes, but also have a role in pathological processes involving the female reproductive tract

Reactive Oxygen Species (ROS) and Male Fertility

Oxidative energy production is inevitably associated with the generation of reactive oxygen species (ROS), excessive concentrations of which can lead to cellular pathology. A free radical may be defined as any molecule that has one or more unpaired electrons. The superoxide anion, the hydroxyl radical, and the hypochlorite radical are some of the highest reactive radicals of oxygen. Owing to their high reactivity and to their capability of initiating an uncontrolled cascade of chain reactions, ROS produce extensive protein damage and cytoskeletal modifications and inhibit cellular mechanisms. Aerobic organisms are equipped with a powerful battery of mechanisms that protect them from the adverse effects of lipid peroxidation (LPO) and other manifestations of oxygen toxicity. Defective sperm function frequently causes male infertility, due to abnormal flagella movement, failure to recognize the zona, and inhibition of sperm-oocyte fusion. ROS are fundamental mediators of physiological sperm function, such as signal transduction mechanisms that have an effect on fertility. ROS can have positive effects on sperm and the concentration functions depending on the nature and the concentration of the ROS involved. They are necessary in regulating the hyperactivation and the ability of the spermatozoa to undergo acrosome reaction. An increased amount of superoxide anion (O2-) is one of the first steps required by the spermatozoa for induction and development of hyperactivation and capacitation. Numerous studies have shown that oxidative stress plays an important role in the pathophysiology of infertility and assisted fertility. The paternal genome is of primary importance in the normal embryo and fetal development. ROS-induced sperm damage during sperm translation, such as signal transduction through the seminiferous tubules and epididymis, is one of the most important mechanisms leading to sperm DNA damage. Male germ cells are extremely vulnerable to oxidative stress as the sperm membrane is rich in unsaturated fatty acids and lacks the capacity for DNA repair. Spermatozoa are particularly susceptible to ROS-induced damage because their plasma membranes contain large quantities of polyunsaturated fatty acids (PUFA) and their cytoplasm contains low concentrations of the scavenging enzymes. Many clinical and research institutes are investigating the usefulness of antioxidant supplementation and their role in prevention of the infertility problems. Incubation under oxygen in vitro was detrimental to human spermatozoa, decreasing motility and viability. Since then, many reports have associated ROS with impaired sperm function, including decreased motility, abnormal morphology, and decreased sperm-egg penetration. Increasing knowledge of the mechanisms whereby ROS and endogenous antioxidant systems influence reproductive processes can assist to optimize the application of exogenous antioxidants to fertility treatment

Redoxomics and Oxidative Stress: From the Basic Research to the Clinical Practice

Potentially oxidant chemical species, which include not only free radicals but also other oxidizing chemical species such as reactive oxygen species (ROS), for example, hydroxyl radical and hydrogen peroxide, and nitrogen reactive species (RNS), for example, nitric oxide, play a relevant role in all biological processes and especially in cell defenses and molecular signaling. Their action is finely modulated by the antioxidant network that is composed either by endogenous or exogenous compounds (e.g., enzymes, peptides, lipids, and vitamins). An impaired modulation of oxidant species can lead to the so-called oxidative stress that is now considered an emerging health risk factor in almost all living organisms including plants, animals, and humans. Indeed, oxidative stress is related to a reduced lifespan and many diseases (e.g., cardiovascular diseases, neurodegenerative disorders, and metabolic diseases) both in humans and in animals. Unfortunately, oxidative stress does not show any clinical picture, but it can be detected only by means of specific laboratory tests. The recent recognition of a specific “redox code” and the definition of a redoxomics as a new “omics” are now enlarging the horizon of the traditional oxidative stress field leading to the definition of the so-called electrophilic stress. The aim of this chapter is to review the basic principles of redox reaction starting from the concept of free radicals and antioxidant in order to define the “electrophilic stress” as an emerging health risk factor for early aging and almost 1000 illness from infectious diseases to cancer. A paragraph is dedicated to the tests to measure oxidative stress in clinical practice either in humans or in animals in order to prevent, to treat and to monitor electrophilic-related diseases

Salto de Volturno

De los tres aprovechamientos hidroeléctricos correspondientes ala cuenca del rio Volturno (Italia), el denominado n.°2 presenta dos características notables: La primera es la de derivar las aguas del rio, llevarlas con una tubería en carga a la proximidad de un pozo, descendiendo vertical y paralelamente a éste hasta llegar a la turbina instalada en el fondo del mismo, desaguando en galería y, finalmente, restituyendo en el rio. La segunda es la casa central, que cubre el pozo y que constituye en sí un ensayo de tipo arquitectónico

Assisted Reproductive Technologies in Safeguard of Feline Endangered Species

The growth of the human population and the escalating consumption of natural resources have reduced wild habitats, modifying the existing balance of biological cycles. Therefore, ex situ conservation efforts have received renewed attention as a potential safeguard for species with an uncertain future in the wild. Most wild felid species are classified as rare, vulnerable, or endangered due to poaching and habitat loss. Any directed action taken by humans to enhance animal reproduction results in assisted reproductive technologies (ART) development. These technologies have been included in programs for the conservation of endangered species. Therefore, ART provide a new approach in the safeguard programs of felid biodiversity. Currently, ART mainly include Artificial Insemination (AI); In Vitro Embryo Production (IVEP) consisting of In Vitro Maturation (IVM), In Vitro Fertilization (IVF), In Vitro Culture (IVC), Embryo Transfer (ET), and Intra Cytoplasmic Sperm Injection (ICSI); gamete/embryo cryopreservation; gamete/embryo sexing; gamete/embryo micromanipulation; Somatic Cell Nuclear Transfer (SCNT); and genome resource banking

Ultrasonographic appearance of early embryonic mortality in buffalo (Bubalus bubalis)

Embryonic mortality is one of the main causes responsible of the decline in fertility that occurs in buffaloes during periods of increasing daylight length (out sexual breeding season). Transrectal ultrasonography for pregnancy diagnosis offers some advantages over palpation per rectum: earlier diagnosis of pregnancy/non-pregnancy, determination of embryo/fetus viability, reduction of misdiagnosis, and reduction of .potential. iatrogenic embryo/fetal attrition. Non pregnant buffaloes on Day 25 after AI showed higher Resistive Index (RI) (P<0.05) and Pulsatility Index (P=0.07) values, registered on CL on Days 10 after AI, compared to pregnant buffaloes. RI values were significantly higher (P=0.02) in non pregnant buffaloes also on Day 45 after AI. Colour Doppler sonography could be used to gain specific information relating to the ovarian blood flow in predicting early embryonic loss and to describe the ultrasonographic features of early embryonic death in buffaloes

The HELLAS2XMM survey. IX. Spectroscopic identification of super-EROs hosting AGNs

We present VLT near-IR spectroscopic observations of three X-ray sources characterized by extremely high X-ray-to-optical ratios (X/O>40), extremely red colors (6.3<R-K<7.4, i.e. EROs) and bright infrared magnitudes (17.6<K<18.3). These objects are very faint in the optical, making their spectroscopic identification extremely challenging. Instead, our near-IR spectroscopic observations have been successful in identifying the redshift of two of them (z=2.08 and z=1.35), and tentatively even of the third one (z=2.13). When combined with the X-ray properties, our results clearly indicate that all these objects host obscured QSOs (4e44 < L(2-10keV) < 1.5e45 erg/s, 2e22 < N_H < 4e23 cm-2) at high redshift. The only object with unresolved morphology in the K band shows broad Halpha emission, but not broad Hbeta, implying a type 1.9 AGN classification. The other two objects are resolved and dominated by the host galaxy light in the K band, and appear relatively quiescent: one of them has a LINER-like emission line spectrum and the other presents only a single, weak emission line which we tentatively identify with Halpha. The galaxy luminosities for the latter two objects are an order of magnitude brighter than typical local L* galaxies and the derived stellar masses are well in excess of 10^11 Msun. For these objects we estimate black hole masses higher than 10^9 Msun and we infer that they are radiating at Eddington ratios L/L_Edd < 0.1. We discuss the implications of these findings for the coevolution of galaxies and black hole growth. Our results provide further support that X-ray sources with high X/O ratios and very red colors tend to host obscured QSO in very massive galaxies at high redshift.Comment: 7 pages, 5 figures, accepted for publication in A&