Process Essay, 18 pages (4500 words)

Steps in the process of fertilization

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Fertilization is an essential process in sexual reproduction which involves the union of two specialized cells called the gametes to form a zygote. This then develops to form the new organism. During the process of fertilization innumerable number of sperms hit the surface of the eggs. But only one sperm succeeds in fertilizing it and the rest are repelled from the surface of the egg. This is done by the modification of cell envelope & extracellular matrix besides altering the metabolic activity of the zygote or embryo. Various metabolic changes occur after the zygote is formed. This includes the respiratory burst hydrogen peroxide production activation of oxidases peroxidase alterations in the redox status changes in the activity of redox-sensitive transcription factors etc. Besides these an embryo-derived paf (1-o-alkyl-2-acetyl-sn-gylcero-3-phosphocholine) is synthesized and its release involves the binding to extracellular albumin which protects its enzymatic degradation (ONeill C, 1985). The change in the redox state affects the activity of redox-sensitive transcription factors that may alter gene expression patterns. Besides, this change in the metabolic status also, is responsible for the spatial differences in cell activity especially after compaction and major embryonic events such as fertilization genome activation and cellular differentiation (AJ Harvey et al, 2002). Multiple variations that occurs during this stage are found throughout phylogeny (Wessel G. M. et al., 2001; Shapiro B. M. et al, 1989).

Three discrete steps are involved with the formation of extracellular barrier during the post-fertilisation stage (i. e. after zygote formation):

Following cortical granule exocytosis an autoactivating serine protease separates plasma membrane attachments to the eggs vitelline layer facilitating the separation of this matrix from the egg surface (Haley S. A. and Wessel G. M., 1999).

The structural components of the fertilization envelope (FE) primarily derived from the cortical granules (Wessel G. M. et al, 2001) self assemble into the vitelline layer network and form a distinct pattern of cytoskeleton like structures. (Chandler D. E. & Heuser J. 1980).

The cortical granule-derived enzyme ovoperoxidase is targeted to the FE by the tethering protein proteoliaisin (Somers C. E. et al, 1989).

The structural proteins self-polymerise into fibers. This is followed by an increase in the ovoperoxidase activity in response to alkalization (Deits T. L. and Shapiro B. M., 1986) and an increase in the synthesis of hydrogen peroxide (Foerder C. A. et al 1978; Heinecke J. W. & Shapiro B. M., 1989).

Studies on nutrient uptake during fertilization have been done in various species(Leese & Barton, 1984, Leese, 1991, Rieger, 1992, Rieger et al., 1992, Rieger & Loskutoff, 1994, Gardner, 1999, Gopichandran & Leese, 2003). However uptake of O2 is the major parameter that provides the required indication of overall metabolic status of a single zygote (Leese, 2003) as the production of ATP by oxidative phosphorylation a reaction in which O2 plays a major role (Thompson et al., 1996 Thompson, 2000 Leese, 2003). Furthermore the respiratory rates of single embryos is directly in correlation with quality (Abe & Hoshi, 2003) and with survival following its transfer (Overström, 1992).

When the production of the harmful reacting species eg., destructive oxygen species supersedes the body’s handling capacity through antioxidants, cellular damage occurs. This type of damage is the usual reason for most of the pathological states in animals, especially in nearly half of the infertile men. ROS bring their damage through various routes; the membrane of the sperm is damaged, which causes the motility rate of sperm to reduce and subsequently its inability to fuse with the egg during the fertilization process. ROS also alter the DNA of the sperm, leading to the improper genetic material getting transferred to the next generation. In spite of this, there is an inverse correlelation between the sperms’a ability to produce ROS and their maturation. In the middle of the process of spermatogenesis, the cytoplasm of the sperm is lost due to its compaction (condensation) which is required for the elongation of sperm. This is witness from the study that immature teratozoospermic sperms are featured by the presence of increased residues in cytoplasm in the mid-piece. Besides the cytotoxic damage caused by the levels of ROS in spermatozoa, hazardous oxygen metabolites produced by the leucocytes present in the ejaculate also damages it. This damage is more significant in the assisted conception therapy, where the contamination of the sperms washed, is presumed to the predominant factor determining the success rate of the fertilization, invitro.

The so called reactive oxygen species ROS viz. H2O2 O2-· OH· etc affect the gametes and early reproductive events. ROS, produced by the peroxidation of the lipids, affects and changes the mitochondrial metabolism, besides producing more ROS. ROS are mainly known for their deleterious effects on spermatozoa and hence on male infertility (de Lamirande E et al, 1997, Sharma, RK & Agarwal A, 1996, Shen, H & Ong, C, 2000). Increased production of ROS has been associated with defects in the morphology of sperm (Aziz N et al 2004) inhibition of sperm motility (Armstrong JS et al, 1999, Parinaud J et al, 1997) fragmentation of sperm DNA (Donnelly ET et al, 1999) and premature capacitation (Villegas J et al, 2003). Further ROS also decreases the capacity for sperm-oocyte fusion efficiency and greatly inhibit the in vitro development of the embryo (Johnson MH et al, 1994, Guerin P et al, 2001, Mammoto A et al, 1996).

More than six decades ago, it was discovered that the oxygen radicals (ROS) may have involved in the reproduction of human, especially men. (MacLeod, 1943). But the same was not studied to that extent in the case of female reproductive function, with only little works on pathological and physiological processes. Paszkowski observed that the selenium dependent glutathione peroxidase (SeGPx) was decreased in follicular fluids of women with no record of infertility (Paszkowski et al., 1995; Paszowski and Clarke, 1996). He also demonstrated that the levels of SeGPx were higher in those follicles which yielded oocytes that were successfully fertilized, compared to those follicles which yielded oocytes that failed to fertilize. Increased levels of hydrogen peroxide was found in the unfertilized oocytes & also in fragmented embryos ( Yang et al., 1998). Whereas an increase in the antioxidant consumption was reported by Paszkowski and Clarke (1996) revealing an increase in the ROS activity, when poor quality embryos were incubated. Attaran et al (2000) observed a beneficial role of ROS, with its levels were high in the follicular fluid in IVF conception cycles compared to that of non-conception. Besides acting on sperm or oocytes separately, ROS were also reported to have its deleterious effect on sperm-oocyte fusion also. Studies reporting the fact the decrease in the levels of enzymes like catalase resulting in the loss of sperm motility. Many other studies produced a conclusive evidence for the production of ROS by human spermatozoa and showed that there was indeed an increase in the activity of ROS, in infertile men. If conventional method of invitro fertilization procedures were employed, even the spermatozoa whose DNA is damaged due to ROS, was able to fertilize the oocytes. Velocity measurements in the sperm motility studied under the impact of the ROS (generated by incubation with hydrogen peroxide) showed that the quality of sperm movement was significantly affected, but were motile, extremely; only the percentage motility was affected.

Increased reactive oxygen species production was observed at 7 h and then at 24 h after IVF just before the first cleavage of the embryo. Increased oxidative activity and redox changes at the time of fertilization have been suggested to signal Ca2+ flux after the penetration of sperm. H2O2 besides being the substrate for ovoperoxidase is produced by a calcium-depending mechanism involving the reduction of one molecule of oxygen and the oxidation of two proton donors.

Contrarily low levels of ROS has a positive effect on sperm functions (Bilodeau JF et al, 2000, de Lamirande E et al, 2003) binding of sperm to zona pellucida (Aitken RJ et al, 1989) and the development of embryo in bovine and other mammals (Harvey AJ et al, 2002 Guo Y et al, 2004, Harvey AJ et al, 2004). The concentration of ROS in both intracellular and extracellular are carefully regulated by enzymatic and nonenzymatic mechanisms and also by the presence of a detailed antioxidant defense mechanism in bovine oviductal tissues and fluids (Lapointe J et al 2003). Antioxidant genes especially a few glutathione peroxidases like GPx-1 Gpx-2 and Gpx-3 were differentially expressed along the oviduct. The major enzymes that are capable of metabolizing hydrogen peroxide (H2O2) belong to the family of GPx as well as the oviductal-catalase (Lapointe S et al, 1998, Brigelius-Flohe R, 1999).

The concentration of oxygen that has to be utilized during the culture of embryos, influences the development and quality of the embryos. When the concentration of oxygen was reduced in the culture of mouse embryos, it altered the embryonic gene expression during the post-compaction stage. This has severe consequences on the fetal development of the mouse. (Deanne Feil, 2006). Furthermore blastocysts cultured under decreased O2 tension correlate more closely with in-vivo-recovered blastocysts than in vitro blastocysts cultured under normal O2 tension (Dumoulin JCet al, 1999; Yuan YQ et al, 2003; Johnson MH et al, 1994; Guerin P et al, 2001).

The driving force for changes in the metabolic status of the zygote is the secretion of the cortical granules at least in sea urchins. The sea urchin embryo generates large amounts of H2O2 at the beginning of development of zygote and its levels are meticulously regulated to prevent any possible toxic effect. Because of this the sea urchin system provides greater insights into the control of reactive oxidants in biological systems.

ROS can either positively or negatively affect the reproductive events in vitro. In the fertilization process the oviducts are the site of important processes that occur prior to implantation such as the maturation of oocyte in the initial stages of embryonic development (Harvey AJ et al 2002). They regulate the ROS levels to provide a proper environment for the gametes followed by their fertilization and the subsequent stages in the development of embryo. Vitamin E protects against the loss of the motility of the sperm through the peroxidation of lipids. Hence, supplementation of the same improved the motility of the sperm and enhanced the possibility of fertilization in asthenospermic invidicuals, even when the original sperm motility measured was only 20%.

Spermatozoa exposed to PUFA showed an increase in its oxidative stress (Aitken, RJ, 2006). In particular, the oxidation of DHA- docosahexaenoic acid bound to phospholipid was shown to be one of the predominant factors that controls the mobility of the sperm in vitro. There is a marked cell to cell differences in the life span of sperm samples, which reflects in their susceptibility to lipid peroxidation.

It is a well established fact that when a spermatozoa is subjected to oxidative stress, its membrane and the DNA is damaged through the membrane lipid peroxidation. The probability of this type of damaged spermatozoa will be able to fertilise the egg depends on the rate at which the functions of the sperm is lost. Till date, the research evidence shows that the motility of the sperm, its capacity to enter into acrosome reaction, its ability to integrate with vitelline membrane of the oocytes are all is prone to get affected by the oxidative stress. Of all the ROS, it is the superoxide anion and hydrogen peroxide that causes the deleterious damage to sperm capacitation and hence are the key mediators. The former contributes to the hyperactivated motility of the sperm, while the later is found to be associated with the tyrosine phosphorylation events in sperm capacitation. Current research have shown that the spermatozoa that are subjected to extreme oxidative stress using the method described by Aitken RJ etal (1998) i. e, exposure to hydrogen peroxide and NADPH, have the capacity to reach the normal rate of fertilization with ICSI (intracytoplasmic sperm injection).

The original postulate of the “ free radical hypothesis” was that the ROS led to non specific modification of various biomolecules, such as proteins, lipids and nucleic acids. This is responsible for the etiology of the pathological condition that arises after that. The existence of oxidase activity was first documented while measuring oxygen consumption following fertilization (Warburg O., 1908). The increase in the oxygen consumption upon fertilization of sea urchin eggs is cyanide insensitive and produces H2O2 as the substrate for ovoperoxidase which crosslink the protective FE (Foerder C. A. et al, 1978). The assembly of ovoperoxidase into the fertilization envelope and the cross linking reaction are carefully regulated events that take place in ten minutes following gamete fusion (Weidman P. J. et al 1985).

High oxygen concentrations are deleterious to early mammalian embryonic development (Thompson JG et al., 1990). Heinecke & Shapiro have characterized an oxidase from unfertilized Stronglylocentrolus purpuratus eggs and had demonstrated its role as respiratory burst oxidase of fertilization. Their oxidase appears to be regulated by a protein kinase. It produces H2O2 when stimulated with Ca2+ and ATP and utilizes NADPH but not NADH as a source of reducing equivalents (Jay. W. Heinecke & Bennett M. Shapiro, 1989; Li J Foote RH., 1993; Dumoulin JC et al., 1999).

Some of the transcription factors including PEBP2 AP-1 p53 and NF-κB are known to be regulated by the changes in the redox status and this regulation has been shown to occur through conserved cysteine residues in the DNA-binding regions of these proteins (Hirota K et al, 1997; Hirota K et al, 1999; Ueno M et al., 1999; Akamatsu Y et al., 1997).

Role of ω-3 fatty acids in fertilization

Fatty acids are classified as: saturated monounsaturated and polyunsaturated (PUFA). There are two main classes of PUFA: n-3 (omega-3) and n-6 (omega-6); distinguished by the location of the first double bond i, e., from the three or six carbon from the CH3 (methyl) end of the fatty acid. ω-3 fatty acids are polyunsaturated fatty acids. Examples include α-linolenic acid (ALA) eicosapentaenoic acid(EPA) and docosahexaenoic acid (DHA). The n-3 PUFAs are generated from ALA, found mostly in the chloroplasts of green plants and grass. These essential fatty acids can be converted to longer chain PUFAs, in liver, by desaturation and elongation enzymes, that is common to both. Fatty acid desaturase 2 (FADS2) is rate limiting (Gurr MI et al, 2002). The human cannot synthesize this, because they do not possess the required fatty acid desaturase enzymes and therefore have to be supplemented through diet.

When the gene expression of this FADS2 enzyme is deleted the first step in the PUFA biosynthesis is stopped. This lack of PUFAs and eicosanoids did not interefere with viability or lifespan of female and male fads2-l mice, but resulted in sterility. Wilhelm S. etal (2008) demonstrated that phospholipids substituted with PUFA have a significant role in Sertolic cell polarity and blood-testis barrier, besides the gap junction network between the ovarian follicle’s granulose cells. They are implicated in various process in human, including reproduction, vision, neural development and growth (Gurr MI et al, 2002). For over a long period, these FAs have been implicated in the different stages of vertebrate fertilization. Either of n-6 or n-3 or both influence the reproductive processes through a heterogenous mechanisms, ranging from the providing the precursors for prostaglandin synthesis, steroid hormone biosynthesis, regulating the transcription factors involved.

In ruminants, the pregnancy is established through the ovulation of the eligible oocytes, insemination at the right time and an adequate dosage of estradion and progesterone, during the luteal and follicular stages of the estrous cycle. Oocytes of cattle, when exposed to methyl palmoxirate to prevent the FA oxidation displayed low level of capcity to form blastocysts after fertilization. Moreover, the embryo have to develop completely to prevent luteolysis, which may occur by the interfereons production to inhibit up-regulation of the endometrial oxytocin receptors. Polyunsaturated fatty acids are also reported to modulate the function of the certain transcription factors that controls the gene expression and thus have a effect on the IC concentrations of the enzymes involved in the regulation of PG and steroid hormones synthesis. In bovine endometrial stromal cells and in lutenised granulose cells, both the omega-3 and 6 PUFAs are found to activate the protein kinase C, which activates the phosphodiesterase by phosphorylation. All these processes are affected by the dietary supplementationof PUFAs. Therefore changes in the PUFA sources subsequently reflects in the omega-3 and omega-6 content of the sperm. And these PUFAs are more susceptible to attack by the ROS as reported by various researchers.

A schematic diagram showing a proposed mechanism in which the PUFAs generating the oxidative stress in human spermatozoa.

High levels of poly unsaturated fatty acids in the spermatozoa of infertile human triggers the production of the ROS from a non mitochondrial source, (may be throughthe NADPH oxidase, NOX 5, influenzed by the calcium). Increased reactive oxygen species, ROS, then induces the peroxidation of lipids, which in turn shoots out the phospholipase A2, culminating in the release of more polysaturated fatty acids with subsequent generation of more ROS to perpetuate the oxidative stress.

Dietary supplementation of n-3 PUFAs influence various aspects of fertility starting from conception and throughout the duration of pregnancy. (ESHRE Capri Workshop Group, 2006, Kind KL, et al, 2006). These n-3 PUFAs are incorporated in the phospholipids of the cell membrane and have their effects on membrane composition & function, (Hong MY, 2002), ROS production (Hong MY, 2002 , Watkins SM, 1998), membrane lipid peroxidation (Hawkins RA etal, 1998), regulators of transcription & translation (Narayana BA et al 2001, Davidson LA2004), production of eicosanoids (Chapkins RS, 1991) and IC signal transduction (Ma D, 2004). Lupton JR (2004) had observed that these mentioned actions facilitate the n-3 PUFA-induced suppression of colon cancer. Similarly, alterations in the PG synthesis (mediated through the manipulation of the n-3 PUFAs) has profound effect on fertility, since PGs affect many aspects of fertilization, e. g., ovulation). Dietary supplementation of varying PUFA content to female cattle and to other mammals have found to alter the size and the number of ovarian follicles, the ovulation rate, the production of progesterone hormone by corpus luteum, the length of gestation and luteolysis timings. In male mammals, dietary PUFAs has demonstrated effect on sperm membrane PL composition and on the ability to fertilize (Abayasekara & Wathes, 1999). For instance, sheep fed with diets containing high PUFAs delayed parturition (Baguma-Nibasheka et al. 1999). They also increased the incidence of placenta getting retained in cattle (Barnouin & Chassagne 1991). High supplementation of ALA in diet, during the post partum period improved pregnancy rate in cattle (Kassa et al. 2002). Embryonic mortality was reduced through the suppression of uterine synthesis of prostaglandin F2alpha in cattle was brought about by altering the FA profile in their diets. This strategy of altering the FA profile in the diets, may be used to improve animal productivity by integrating the nutrition and reproductive management (Mattos R etal 2000).

The fatty acid composition of both oocytes and sperm are responsible for the various observations in the study of the fertilization process in animals. The FA composition of the oocytes is specific for a given species both in terms of their abundance and their utilization. Among them, EPA, DHA and ARA have been associated in several stages of reproduction (Wathes DC etal 2007). Mature zebrafish oocytes fortify with ARA, indicating their capacity to synthesize eicosanoids for ovulation and follicular maturation. ARA along with LA was the most predominant PUFAs in oocytes of pig, sheep and cattle. (McEvoy etal 2000). Decreased ARA: EPA ratio in eggs and ovaries of fish have been reported as the reason for poor productivity in the captive broodstock (Pickova J, 2007, Cejas JR et al 2003). DHA along with EPA inhibited the gonadotrophin-mediated steroid hormone synthesis in cold and warm water fishes. These two highly unsaturated fatty acids (HUFA) act as regulator molecules in the maturation of those fish ovary (Mercure F et al 1995).

Kim et al (2001) had observed that the quality of oocytes was influenced by the dietary FAs which changes the composition of granulose cells and oocytes. Based on those quality, the oocytes were divided in to grade1, 2, 3 etc. Kim also observed that there were differences in the FA composition between those grades of sheep’s oocytes, which affected the oocytes competence, resulting in alterations in fertilization rate and developmental potential. Sheep fed with PUFAs had altered FA composition in their membrane PLs of cumulus cells, resulting in the alteration of the oocytes membrane properties (Zeron et al., 2002). Supplementation of n-6 and n-3 showed different reponses in reproduction (Wonnacot KE et al, 2010). Ewes fed with n-3 PUFAs prior to ovarian stimulation and follicular aspiration showed no changes in the follicle number and size. But follicular-fluid levels of progesterone were found to be greater and all the blastocycts (both treated and untreated with the stipulated diet) contained increased levels of PUFAs, mostly ALA. Moreover, dietary supplementation of conjugated linoleic acid (CLA) to early-lactation dairy cows showed an improvement in their reproductive performance (MJ de Veth 2009). Cows fed with 18: 3 FAs rich diets had a prolonged preovulatory follicle at insemination and larger volume of corpus luteum compared to those fed with MUFAs (Bilby TR, 2006).

The deficiency of essential fatty acids affects the energy and fat metabolism, biosynthesis of PUFA, structure of cell membrane and signaling pathways in lipid resulting in incompatible life (Cunnane, 2003). Studies on fads2-/- mouse model had thrown light on the various effects of the deficiency of PUFAs, invididually,( ie, omega-3, omega-6 etc) and combined. In that study, FADS2 deficiency caused hypogonadism and sterility of male (azoospermia) & female mice. Spermatogenesis is stopped in male fads2-/- mice occurred at the stage of round spermatids, leading to azoospermia. This is often caused by a damaged blood testis barrier BTB. BTB is formed by TJ and AJ protein complexes that are restricted to basolateral compartment of the more polarized sertoli cells (Fanning et al, 1998; Chapin et al, 2001; Ebnet et al, 2003). Similarly Thangavelu G etal (2007) have shown that the development of embryo was increased in Holstein cows that are fed with unsaturated fatty acids compared to those with saturated fatty acids. This shows that a diet rich in α-linoleic acid facilitates the development of embryo when compared to that of linoleic or saturated fatty acids. Another observation which showed that, enriching the semen diluents with DHAs containing egg yolk prior to freezing did not enhance the sperm quality following thawing.

In both human and in domestic animals, levels of the breakdown products of lipid peroxidation, such as malondialdehyde and 4-OHalkenal, had no correlation with the semen quality. But, studies on chickens showed that dietary supplementation with more PUFAs, decreased the antioxidant status and semen quality (i. e, sperm concentration and volume). In this context, the significance of the role of the non-enzymic antioxidant, vitamin-E (a chain breaking antioxidant), which reverses the negative effects caused by the PUFA supplementation is to be noted.

Besides all the sources that had been mentioned in this introduction, the final source of oxidative stress is the sperm itself. Defective human sperms produce ROS, which is directly proportional to the extent of the impairment of the sperm function (Deluliis 2006). Another reason for sperm being a source for ROS is that some authors believe that they contain more amounts of unsaturated fatty acids, mainly DHA and AA (Ollero M, 2000). Their study demonstrated that exposing the human spermatozoa to various PUFAs resulted in the accelerated production of free radicals, subsequent to peroxidation of lipids and DNA damage. There is an abnormal retention of remnants of cytoplasm and a presence of increased levels of unsaturated fatty acids in the immature or defective human spermatozoa, which generated high levels of reactive oxygen species. PLA2 gets activated due to this peroxidation of lipid, which enhances the production of more free poly unsaturated fatty acid from the phospholipid further increasing the production of ROS. In this situation, the fertilizing potential of spermatozoa under increased oxidative stress with concomitant low antioxidant status and more ROs production, will decrease dramatically as observed various researchers and in various species.

Dietary n-3 PUFA affects reproductive processes including ovulation & oocyte development and sperm levels & motility. The in vivo morphology of oocytes is improved through a high supplementation of n-3 PUFA (zeron Y, 2002). PUFAs stimulate the generation of in vitro ROS (Aitken RJ et al 2006). In males the fatty acid composition of the sperm membrane influences their fertility and fertile men have much higher sperm levels of omega-3 FAs as compared to infertile men. Infertility and premature birth are two womens health issues where omega-3 levels are implicated. omega-3 supplementation decreased the clotting in the endometrial cells of the uterus and improved the implantation rates of fertilized eggs. Lower concentrations of spermatozoon DHA in asthenozoospermic men are not due to diet but to some type of metabolic difference (Conquer JA). Fouladi et al (2010) have shown that the ovary regulates the effects of alterations in plasma n-3 and n-6 FAs, resulting in only small effects on their developmental potential. The cessation of growth and some health problems in growing rats fed with low PUFAs were reversed after feeding the same with high PUFA sources rich in 18: 3 ALA (Burr and Burr 1930). Reproduction in cattle is influenced more by the type of fats (ie. PUFA or MUFA) than with just fats as it is, reveals the importance of PUFAs in reproductive processes. This is more highlighted because ruminants extensively hydrogenate PUFAs, thereby limiting their supply for absorption in small intestine.

Eicosanoids-independent mechanisms such as modulation of intracellular signaling pathways transcription factor activity and altered gene expression (Das UN., 2000; Dentin R et al, 2005; Simopoulos AP. et al, 2002). Fatty acids in the oocytes are utilized during its maturation and are incorporated into its cytoplasm (Ferguson EM et al, 2006; Kim JY et al, 2001). Changes in the n-3 PUFA levels in the diet alter the fatty acid composition of the oocytes and its surrounding environment affecting the oocytes maturation; modulate the development of follicles, ovulation, embryo development and developmental competence such as its ability to involve in fertilization. The changes in the diet profile for the n-3 PUFAs also had altered mitochondrial properties and increased the ROS levels in oocytes, suggesting a role for mitochondria in the impaired embryo development. Sarah et al (2008) studied the effect of diet supplementation of n-3 PUFA on the zygotes. Exposing the reproductive tract for a period of 22h (post hCG) found to increase the number of zygotes which are morphologically poor, especially when females were given a diet rich in n-3 PUFA. The in vivo-derived zygotes (which were morphologically normal) which were subjected to n-3 PUFA treatment failed to cleave and their development was delayed (Sarah et al, 2008). This was due to the impaired mitochondrial metabolism.

Low and high levels of omega-3 concentrations in the blood have been implicated in various conditions. Hong et al. (2002) have observed n-3 FAs increase apoptosis in colonocytes when coincubated with butyrate. Eicosopentanoicacid increases oxidative stress leading to lipid peroxidation in Walker 256 rat tumor cells besides decreasing the mitochondrial membrane potential (Colquhoun A et al 2001.). in oocytes this change in mitochondrial membrane potential is observed to be a consequence of metabolic inhibitors (Van Blerkom J et al 2003) and have been correlated with developmental arrest in mouse two-cell embryo increased fragmentation (Acton BM et al 2004) and the rate of embryo development in the human (Wilding M et al 2001).

Low omega-3 FA in blood leads to increase in menstrual pain. Lipids of the Oocyte are crucial for the energy requirement of the preimplantation zygote. They also participate in fertilization and in the subsequent cell differentiation (Amri et al., 1994). Oocytes that are oxidatively damaged have a low levels of PUFA and are unable to undergo fertilization (Tarı´n et al., 1996).

The levels of arachidonic acid and docosahexaenoic acid as well as lipid peroxidation in blood and seminal plasma of normozoospermic males from infertile couples compared with that of fertile volunteers indicate that systemic oxidative stress resulting in increased lipid peroxidation and an alteration in the fatty acid profile which may be responsible for infertility in men (Oborna I et al, 2009).

The omega-6 or -3 fatty acids are the precursors for various metabolites produced in sperm and ovum which are necessary for fertilization. The precursors from omega-3 are less potent in generating ROS than omega-6. Therefore the gametes will be less affected by the effects of ROS under this condition. An imbalance in the omega-6-to-omega-3 ratio has been linked to various complications; like polycystic ovarian syndrome PCOS low sperm count etc.

Treatment of cumulus-oocyte complexes (COCs) with ALA significantly increased the percentage of oocytes at the metaphase II an increase in the percentage of cleaved embryos the blastocyst rate and better -quality embryo compared with untreated controls while higher doses of it were detrimental (Waleed F et al, 2009). Thus the omega-3 FAs and the Redox regulators have multifarious roles before during and after the fertilization process.

The role of the polyunsaturated fatty acids is important in both the physiology of sperm and ovum. This is highlighted by their key role in the maintaining the fluidity of the membrane of the sperm, which is needed for fertilization. Further, they are also a important part of the specific class of fucosylated slycosphigolipids, which are important for the male fertility. Besides these, they are also sources of alkoxyl and peroxyl radicals, to help these cells in the event of damages arising due to oxidative stress.

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