Justification and PCOS and Fertility - Thesis Example

1.0 INTRODUCTION 1.1 Infertility Marques-Pinto & Carvalho (2013, p.15) define infertility as the inability to conceive after one year of unprotected sex. The prevalence of infertility worldwide is 9% and it is estimated that in infertile couples, the cause is primarily feminine in 38 percent and largely masculine in 20 percent. WHO defines infertility as the disease of the reproductive system manifested by the failure to attain clinical pregnancy after 12 months or more of regular unprotected sex (WHO, 2015). Brugo-Olmedo et al (2000) also defines infertility as the inability of a sexually active couple not under contraceptives to attain pregnancy within a period of one year. The male partner can be assessed for infertility using various clinical interventions or by evaluating semen in the laboratory. Demographic definition of infertility include the inability of individuals in the reproductive age of (15-49 years) to become pregnant or complete pregnancy to the term within 5 years of pregnancy exposure. Another demographic definition of infertility is the inability to become pregnant with a live birth in 5 years of consistent sexual intercourse without using any contraceptives (WHO, 2015). According to WHO (2010), there are two forms of infertility, namely primary infertility and secondary infertility. Primary infertility refers to the inability of a woman to ever bear children, either because of being unable to get pregnant or not being able to carry pregnancy to a live birth. Therefore, women who have spontaneous miscarriage or who have still births without ever having had a live birth, suffer from primary infertility. On the other hand, secondary infertility refers to a woman not being able to bear children, either because they are not able to become pregnant or carry pregnancy to a live birth after having had a previous pregnancy. Therefore, women who constantly have spontaneous miscarriages or have stillbirth after previous pregnancy or ability to conceive and later on are not able to carry pregnancy to live birth manifest primary infertility. Male infertility is also an important factor. According to Kumar & Singh (2015, p.192) male infertility is the inability of a male to impregnate a fertile female. Male facto infertility is perceived as the change in sperm concentration in a sperm sample, collected one and four weeks apart. In human beings, sperm concentration accounts for 40-50 percent of infertility and affects about 7 percent of all men. Male infertility is mostly caused by semen deficiencies as well as low semen quality. Kumar & Singh (2015, p.192) further explain that males with sperm parameters lower than the WHO normal values are perceived to have male factor infertility. The common aspects in regards to male infertility include; low sperm concentration, poor sperm motility, and also abnormal sperm morphology. Semen volume, as well as other seminal markers of epididymal, prostatic, and seminal vesicle function; are some of the factors allied to male infertility (Kumar & Singh 2015, p.192). According to Kumar & Singh (2015, p.192) about 90 percent of male infertility is associated to sperm count and there is a positive correlation between the abnormal semen limits and sperm count. Problems with sperm count, sperm motility as well as sperm morphology is as a result of disorder in control mechanism, as well as pre-testicular, testicular, and post-testicular factor (Kumar & Singh, 2015, p.192). 1.2 Justification and PCOS and Fertility Infertility is defined as the inability to conceive after a reasonable period of sexual intercourse without any contraceptives measures. Infertility has serious consequences that include serious economical impacts and burdensome problems on the affected couples. Additionally, infertility has also been associated with serious consequences such as marital conflicts, domestic violence, stigmatization, isolation and as well as separation and divorce. Infertility has been attributed to many factors and polycystic ovarian syndrome (PCOS) is among these factors. According to Amer (2009, p.263) PCOS is the most common cause of ono-vulatory infertility and accounts for approximately 75 percent of cases. PCOS is a syndrome of ovarian dysfunction allied to hyperandrogenism and polycystic ovary morphology (Amer 2009, p.263). Mostly, fertility problems in women having PCOS are due to the lack of ovulation, although absence of ovulation might not be the only cause of infertility. Therefore, it is important to understand mechanisms that control development of oocytes and ovulation because lack of ovulation is the main cause of PCOS and infertility. Understanding these mechanisms is important because it can aid in developing effective assisted reproductive technologies. 1.3 Ovarian follicle Ovaries are the main organ within the female’s reproductive organ and they contain egg cells. The egg cells are located within the follicles and after the onset of puberty; various hormones stimulate maturation of follicles to release egg cell, known as ovum. Hormones play the role of regulating menstrual cycle and also stimulate ovulation. During ovulation, the ovum travels down the fallopian tube into the uterus. Ovulation refers to the moment when the ovum leaves the ovary (Acton, 2013). Ovarian follicles refer to the fundamental units of female reproduction system and every ovarian follicle contains a single oocyte. Ovarian follicles are sporadically instigated to grow and develop, which eventually culminates in ovulation of normally a single element of oocyte. About ten ovarian follicles starts maturation during a usual menstrual cycle and out of these ovarian follicles, one of them develops into a dominant ovarian follicle (Acton, 2013). During ovulation, the primary follicle develops into secondary follicle and eventually turns into mature vesicular follicle. The mature vesicular follicle finally turns into corpus leteum after rapture. Every month, an oocyte which is a mature ovum is released by one of the ovaries and the primary oocyte develops within the follicle. Follicle provides the required steroid and hormones necessary to maintain the reproductive cycle, secondary sexual characteristics as well as provides the necessary hormones to prepare the uterus for implantation (Acton, 2013). 1.4 The reproductive cycle The reproductive cycle refers to the cycle of natural changes occurring within the uterus and ovary and it is fundamental in ensuring sexual reproduction. The reproductive cycle is important for the production of eggs as well as in preparing the uterus for pregnancy. The cycle takes place within fertile females and occurs constantly between onsets of puberty until the end of menopause. The reproductive cycle consists of the ovarian/uterine cycle. The ovarian cycle consists of changes that take place within the follicles of the ovaries while the uterine cycle consists of changes that occur within the endometrial lining of the uterus. The two cycles consist of 3 phases where the ovarian cycle includes the follicular phase, ovulation and the luteal phase. On the other hand, the uterine cycles includes the menstruation, proliferative phase, in addition to secretory phase. The increase of the follicle stimulating hormone stimulates some ovarian follicles during the initial days of the menstrual cycle. Hormones influence stoppage of development of other follicles and only one dominant follicle within the ovary develops to maturity. When the follicle matures, it contains the ovum (Prior et al, 2015). The second phase of the ovarian cycle is ovulation. During this phase, the ovarian follicle releases the mature egg into the oviduct. Following the release of the egg from the ovary, the egg travels to the fallopian tube and in case the egg is not fertilized, it disintegrates within the fallopian tube. The last phase is the luteal phase where the FSH and LH induce the transformation of the remaining parts of the dominant follicle into the corpus leteum and this leads to production of progesterone. Increased levels of progesterone stimulate the production of estrogen. These hormones suppress the production of the FSH and LH required for maintenance of corpus luteum. As a result, the level of FSH and LH drops rapidly and eventually atrophies. The reducing progesterone stimulates menstruation and the onset of uterine cycle (Prior et al, 2015). Menstruation is the first cycle of the uterine cycle and menses indicates that the woman is not pregnant. The second phase is proliferative phase where there is growth of the uterus lining. As the uterine lining matures, the ovarian follicles produce increasing levels of estradiol and estrogen. Estrogens instigate formation of proliferative endometrium as well as induce crypts within the cervix which leads to production of fertile cervical mucus. The last phase is the secretory phase where the corpus luteum produces progesterone. Progesterone plays an important role in preparing endometrium for implantation of the blastocyst. Progesterone also supports early pregnancy by elevating blood flow as well as uterine secretions and decreasing the contraction of the smooth muscle within the uterus (Prior et al, 2015). 1.5 Oestrous cycle Oestrous cycle is a cycle of the reproductive organs that is controlled by hormones. The cycle begins after sexual maturity and is disrupted by anestrous phase or pregnancy. Generally, the cycle continues till death. It includes the follicular stage, ovulation and is ensued by the luteal phase and the regression follows which causes the return to the first phase of the cycle (Satué & Gardón, 2013).   1.6 The menstrual cycle The menstrual cycle starts on the 1st day the blood discharge starts and ends on the day prior to the subsequent menstrual period. The blood discharge takes place because the body is getting rid of the lining that was prepared during the preceding cycle in preparation for pregnancy. The body also rids the egg that was not fertilized during the last cycle. With the new cycle, the body then starts to prepare itself again for egg release and preparing the uterus for pregnancy (Prior et al, 2015). 1.7 Folliculogenesis Folliculogenesis refers to the ovarian follicle development process. For the ovarian follicle to reach the ovulatory phase, the follicle passes four stages, namely: primordial stage; pre-antral stage; antral stage; and lastly the pre-ovulatory follicle stage (Yang et al, 2013, p.213). The focus of the project on female fertility with emphasis on the mechanisms controlling folliculogenesis and normal ovulation and hence it is important to understand the human ovarian folliculogenesis because the understanding folliculogenesis has important implications in assisted reproduction technology and fertility preservation as well. Folliculogenesis is strongly regulated through crosstalk between cell death and survival signals. Folliculogenesis combines a complex programme of synchronised follicle development and oocyte maturation which relies on a myriad of cell signalling entities criss-crossing between follicular compartments (Hsueh et al, 2015, p.3). Diminution of the ovarian follicular reserves begins during fetal life and carries on throughout a female’s life span. However, only a small percentage of the primordial follicles reach the ovulatory phase, whereas the other follicles undergo atresia which is a degenerative process. During a natural menstrual cycle, only one follicle undergoes ovulation whereas other follicles undergo atresia under the correct regulation of hypothalamus-pituitary-ovary axis and intraovarian regulators (Yang et al, 2013, p.213). Nonetheless, within the controlled ovarian stimulation during ART, the hypothalamus-pituitary-ovary is controlled and regulated using medications. For instance, desensitizing and down-regulating the pituitary gland via GnRH-agonist long procedure facilitates intiation of exogenous gonadotropins on the day of anticipated wave emergence. In addition, administering the exogenous gonadotropins can result to more than one selection within a menstrual cycle. Therefore, it is of utmost importance to evaluate the follicular fluid exosomes because this will also provide knowledge and an overview on human ovarian follicular signalling and waves (Hsueh et al, 2015, p.5). 1.8 Cell- cell communication during follicle development According to Battaglia et al (2017, p.1) the mammalian ovarian follicle consists of germ cell, somatic cells and follicular fluid. Paracrine communication among the various types of cells via follicular fluid facilitates successful development of a mature oocyte in readiness for fertilization. During embryonic life, primordial germ cells move though the follicular fluid to the genital ridge through mitosis before transforming into primary oocytes. The primary oocytes undergo meiosis and are arrested within prophase I (Battaglia et al 2017, p.2). During this stage the oocyte is enclosed within ovarian somatic cells and a primordial follicle is formed. Within the antral follicles, the oocytes are stimulated by hormonal signaling and meiosis starts once again. As aforementioned, only single follicle ovulates and releases a mature egg for fertilization. During this phase, the ovarian follicles have the germ cell within the metaphase of the 2nd meiotic division within a fluid-filled cavity as well as different layers of cells (Santonocito et al, 2014, p.1753). There is close relationship between follicle development and oocyte maturation and actually the proliferation and differentiation of somatic follicular cells takes place systematically with the maturing oocyte, mediated through continuous exchange of signals between somatic cells and germ cell. According to Battaglia et al (2017, p.2) the communication between the oocyte and somatic follicular cells takes place though gap-junctions formed between the oocyte and also via the follicular fluid built in within the antrum. Communications within the follicular fluid occurs through autocrine and paracrine mechanisms and the miRNAs within the ovarian follicle mediates the communications (Santonocito et al, 2014, p.1753). According to Battaglia et al (2017, p.2) the miRNAs are vital regulators within the mechanisms involved in folliculogenesis as well as oocyte maturation. As aforementioned, the crosstalk between the oocyte and somatic follicular cells takes place mostly through gap junctions found between the follicular somatic cells. Proteins that form the gap junctions are known as connexins (Cx). Both Cx43 and Cx45 take part in GC communications whereas Cx37 and Cx43 participate within oocyte and CC communications (Pietro 2016, p.302). Studies show that the various kinds of gap junctions within crosstalk between the CC and the oocyte have varying permeability characteristics and therefore have the ability of transferring specific signals and molecules (Corrado et al, 2013, p.2). In addition, the follicular fluid built in within the antral follicle is another means used for communication between somatic cells and oocyte. As Pietro (2016, p.306) further explains, follicle fluid is made up of nucleic acids, metabolites, ions and also proteins. The oocyte, granulose, and theca cells secrets these components, together with plasma components that cross the blood–follicular barrier through theca capillaries. Follicular fluid represents an extremely fundamental micro-environment for the oocyte to develop. Additionally, the biochemical composition of the follicular fluid indicates the physiological condition of the follicles. Therefore, analysis of the components of the follicle fluid can offer valuable information regarding the quality of oocyte. Actually, the follicular cells within the follicular fluid secrete cytokines, hormones, growth factors and chemokines that further promote the maturation of oocyte (Pietro 2016, p.306). On the other hand, modification of the biochemical composition of follicular fluid is associated with low quality oocytes. Therefore, assessment and evaluation of different components within the follicular fluid can assist in understanding intra-follicular communications and signaling and also identification of the potential biomarkers of oocyte health for women going through assisted reproductive treatment (Pietro 2016, p.304). 1.9 Extracellular vesicles Extracellular vesicles are potent modulators of the immune system that play the role in immune signaling during physiological and pathological processes. The extracellular vesicles play the role of modulating the local maternal immune function. In early phases of the human reproductive process, the potential sources of the extracellular vesicles include ovarian follicle, seminal fluid, embryo and endometrium (Tannetta et al, 2014, p.540). During later stages of pregnancy, the STP of the placenta produces the extracellular vesicles. The extracellular vesicles are released into the maternal blood and they act as the signaling mechanism between fetus and mother. Signalling mechanism of the extracellular vesicles occurs via protein or lipid ligand-receptor interactions or micro-interfering RNAs (miRNAs). miRNAs are responsible for regulating gene expression and also extracellular exchange of genetic material, which control immune responses (Tannetta et al, 2014, p.540). 1.10 Follicular fluid Follicular fluid encompasses a composite combination of nucleic acids, proteins, metabolites, as well as ions that oocytes and somatic cells secrete. Studies have demonstrated the presence of microRNAs (miRNAs) within the human follicular fluid and are transported by extracellular vesicle, like microvesicles and exosomes (Battaglia et al 2017, p.2). Follicular fluid offers a micro-environment for the development, maturation and ovulation of oocytes. The follicular fluid has different types of hormones, proteins, metabolites, along with regulatory molecules that significantly help in the development and maturation of oocytes. 1.11 Exosomes Exosomes are the secreted membrane-bound nano-vesicles. Biogenesis of exosomes occurs within the late endosomal membrane partition through inward budding of the restraining membrane of late multivesicular endosomes⁄multivesicular bodies (MVB) (Mincheva-Nilsson & Baranov, 2010, p.522). Exosomes have cell surface–expressed proteins and cytosolic constituents. Secretion of the exosomes occurs within the extracellular space by fusing of the MVB with the plasma membrane. Proteins that are carried by the exosomes are sorted to the MVB through endocytosis and transportation of plasma membrane proteins to the early recycling and further to the late endosomal partition. The other way is by being directly transported from the golgi complexi to the MVB and being inserted within the MVB membrane to sprout as exosomes (Mincheva-Nilsson & Baranov, 2010, p.523). 1.12 miRNAs The miRNAs refer to small non-coding RNAs that suppress gene expression. For the miRNAs to repress gene expression, they target specific mRNAs mostly within their 3’ untranslated area. As a result, they are extremely vital for post-transcriptional gene regulation. In addition, miRNAs are involved in the regulation of the activity of many protein-coding genes (Santonocito et al 2014, p.1). Studies have shown that human follicular fluid contains miRNAs. The miRNAs are normally around 22 nucleotides long and function as small post-transcriptional regulatory molecules that act through binding to their specific mRNA targets. The miRNAs play this role thorough degeneration of mRNAs or hindering them to be translated to proteins. The significance of this study is that even though miRNAs are involved in numerous physiological processes and there have been extensive research within other body fluids, there are few studies regarding miRNAs within the follicular fluid (Santonocito et al 2014, p.1). Feng et al (2015, p.5) found out that some miRNAs responsible for the regulation of steroidogenesis in vitro are allied to polycystic ovarian syndrome (PCOS). Another study conducted by Diez-Fraile et al (2014, p.1) showed that age-related differential miRNAs levels in follicular fluids of human can show pathways potentially establishing fertility and success of in vitro fertilization. This indicates the significance of evaluating miRNAs within the follicular fluid in regard to assisted reproduction as well as reproductive ageing. Santonocito et al (2014, p.6) identified 37 miRNAs up-regulated within the follicular fluids. Analysis of these miRNAs demonstrated that miRNAs participate in vital pathways involved in the development and maturation of oocyte. Therefore, the miRNAs might signify non-invasive molecular markers of the quality of oocyte (Santonocito et al 2014, p.6). Biogenesis of miRNAs (HasanSohel 2016, p.178) 1.13 Role of miRNA in ovarian function - use of knockout studies According to Feng et al (2015, p.6) miRNAs play important role in various signaling pathways. Specifically, miR-320 is involved in regulation of various signaling pathways and this include both Wnt signaling and insulin-P13K signaling. Wnt signaling pathway is involved in the regulation of numerous diverse cellular processes that include cell proliferation, establishing polarity, pluri-potency, among other cellular processes. Studies have shown that expression of elements of Wnt signaling pathway occurs within the ovaries of mouse and also their oocytes. In addition, some studies also show that Mnt signaling pathway is fundamental during pre-implantation embryo development as well as in development of ovarian follicle (Feng et al 2015, p.5). According to Takezawa et al (2011, p.4) during fertilization the family of Wnt signaling pathway participates in both cell membrane adhesion and fusion. Some studies also show that canonical Wnt/β-catenin signaling pathway is involved in the regulation of embryo development to blasto-cyst phase. In addition, according to Feng et al (2015, p.6) specific inhibition of Wnt/β-catenin signaling pathway can encourage blastocyst hatching in pigs and also increase the number of cells. The miR-320 is involved in Wnt signaling pathway and a study conducted by Denicol et al (2011, p.2) demonstrated that activating canonical Wnt signaling pathway following a main zygote genome activation is likely to reduce the bovine embryo development to blastocyst and also decrease cell numbers of inner cell mass. Denicol et al (2011, p.3) further demonstrated that WNT signaling antagonist Dickkopf-1 has the ability to enhance embryo survival following embryo transfer to females during assisted reproduction. Evidence also shows that if the canonical Wnt signaling pathway is inhibited, this can hinder blastocyst ability to implantation (Feng et al, 2015, p.5). Since miR-320 regulates Wnt signaling pathway, there are some studies that have shown that Wnt signaling pathway may impact fertilization as well as early embryo development and it can be distorted through knocking down the level of miR-320 (Feng et al, 2015, p.5). As per Feng et al (2015, p.10) miR-320 and miR-197 in human follicular fluid have correlation with embryonic development potential. This is because in this study it was established that Knocking down miR-320 within mouse oocytes has negative impact on embryonic development potential because this prevents Wnt signaling pathway to be expressed. This study further reflects that miRNAs within follicular fluid have an impact on the quality if the embryo (Sodel et al, 2013, p.6). Furthermore, this provides the basis for the future development of miRNAs within the follicular fluid as protein markers for determining embryo quality as well as other phenotypes associated with fertility. 1.14 Extracellular miRNA These are miRNAs found within the extracellular environment and this includes various biological fluids. Evidence indicates that extracellular miRNAs are also within extracellular fluids like saliva, breast milk, colostrum, peritoneal fluid, as well as seminal fluid (HasanSohel 2016, p.178). In addition, the expression profile of extracellular miRNA from various forms of bio-fluids in regard to various patho-physiological conditions demonstrates a precise pattern that indicates that extracellular miRNAs are selectively released from the cells. Studies show that extracellular miRNA can function as an intracellular communication system within the body (HasanSohel 2016, p.180). References Amer S, 2009, Polycystic ovarian syndrome: diagnosis and management of related infertility, Obstetrics Gynecology Journal, 9(1), pp:263-270. Read More