INTRODUCTION:

The failure of a couple to conceive after 12 months of consistent sexual activity without the use of contraception for women under 35 and after 6 months for women 35 and older is known as infertility ¹. Infertility can be divided into primary infertility or secondary infertility. Infertility can be due to male causes, female causes, or combined and unexplained causes². Infertility in women can be caused by a variety of factors, the most prevalent of which are problems with ovulation, such as polycystic ovarian syndrome ^3-13

Since it serves as a springboard for subsequent embryonic growth and a healthy pregnancy, implantation is an essential stage in mammalian reproduction. The "window of implantation" or "window of receptivity" refers to the brief period of time during which the embryo and the uterus must engage in reciprocal interactions inside it ¹⁴. Any communication breakdown between the embryo and the endometrium during this period might lead to implantation failure. In reproductive medicine, implantation failure is seen as an unresolved issue that results in infertility ¹⁵.

Complex locally active substances released at the implantation site and involved in the early embryo–uterine contact include growth factors, hormones, cytokines, and their cross-talk ¹⁶. These chemicals form a network that helps prepare the receptive endometrium and blastocyst. The quality of the blastocyst, the existence of a receptive endometrium, and the synchronization of the embryo's developmental phases are believed to be necessary for successful implantation. In addition to ovarian dysfunction resistant components, the implantation process is dynamic and involves coordinated autocrine, paracrine, and endocrine effects ¹⁷.

Progesterone is one of several hormones that may affect the implantation process by spatiotemporally mobilizing many molecular modulators ¹⁸. The most crucial hormone for endometrial development is progesterone. Both stromal and epithelial tissues have progesterone receptors, and the amounts of these receptors as well as hormone concentrations are essential for successful implantation ¹⁹. A greater estrogen-to-progesterone ratio has been shown to affect the aspect of integrin molecules, which play a crucial role in blastocyst adhesion to the uterine epithelium ²⁰.

Menstruation, ovulation, and implantation are just a few of the reproductive processes in which prostaglandins (PGs) are known to be important. The cycloxygenases (COX-1 and COX-2) are the main enzymes that produce PGs. According to the precise arrangement of the Cox-1 and Cox-2 genes in the preimplantation uterus, PGs are essential for the implantation of embryos. Implantation failure occurs when PG production is inhibited either prior to or during implantation ²¹.

The transmembrane tyrosine kinase known as the epidermal growth factor receptor (EGFR), which is widely expressed, binds to amphiregulin (AREG). A membrane-anchored precursor protein that may take part in cell-to-cell communication, AREG is created. Alternatively, AREG functions as an autocrine or paracrine factor and is secreted after being proteolytically processed by cell membrane proteases. Growth factors, hormones, cytokines, and inflammatory lipids are some of the triggers that trigger the production and release of the AREG gene. By binding to EGFR, AREG controls crucial intracellular signaling cascades that govern cell motility, proliferation, and survival. In a physiological sense, AREG plays a role in the development and maturation of bone tissue, oocytes, and mammary glands ²².

Ovarian stimulation, ovum collection, fertilization, embryo cleavage, and implantation are all steps in the multi-stage process of assisted reproduction. As the initial step toward a healthy baby, achieving a viable intrauterine pregnancy is the aim of each of these treatments. The quality of all reproductive components, especially embryos, affects the success rate. Therefore, the enhanced selection of growing embryos is one of the most challenging challenges in IVF programs ²³. Therefore, the number of blastomeres, size equality, and fragmentation percent are the morphological criteria used to identify healthy oocytes and high-quality embryos at the 2–8 cell stage ^{24, 25}. There is substantial evidence that numerous reproductive hormones are important for folliculogenesis, oocyte maturation, corpus luteum formation, and endometrial preparation for implantation of the fertilized ovum, and that any disruption in their levels can impact fertility. As a result, little is known about the processes controlling the ovarian follicles' cyclic recruitment, selection, and dominance ²⁶. Follicle cell sensitivity to gonadotropins and other systemic factors can be altered by intra-ovarian variables.

The mature follicle contains both germ cell components (oocyte) and components of somatic cells (mural granulosa cells and cumulus) ²⁷. A basement membrane separates it from the rest of the ovary. During follicular development, the oocyte and somatic cells coordinate their growth²⁸. Within the particular environment of the ovarian follicle, endocrine, autocrine, and paracrine signals govern and coordinate this process ²⁹.

Consequently, the follicular fluid (FF) acts as a channel for the passage of signaling mediators into and out of the follicle, as well as between other cell types inside the follicle³⁰. It is reasonable to assume that certain biochemical characteristics of the FF influence the quality of the oocytes and, consequently, their capacity to fertilize and create embryos ³¹.

MATERIALS AND METHODS:

Patients:

This study was performed on 100 infertile couples, who attended the Kamal AL-Samarai Hospital, the center of fertility and IVF in Baghdad-Iraq. The duration of the study extended from September 2020 to March 2021.

The female samples having intracytoplasmic sperm injection (ICSI) varied in age from 20 to 35 years, with a BMI between 15 and 27 kg/m². Fifty infertile women with polycystic ovarian syndrome (PCOS) and fifty infertile women without PCOS were selected from among the 100 women. Women who were stimulated by extended GnRH agonists were included in each group; those who had uterine fibrosis or endometriosis were not. On the day of ova collection and 14 days after embryo transfer (ET), each patient's serum levels of progesterone (PROG) (ng/ml), prostaglandin (PGE2) (ng/ml), and humen keratinocyte autocrine factor amphiregulin (KAFAREG) (pg/ml) were assessed. The levels of those hormones in follicular fluid were also assessed following ovum harvest.

Materials and hormonal assay kits and procedure

Sample collection and storage:

Blood samples were taken from each woman who had had the ICSI technique. After allowing the blood to coagulate for 30 minutes, the serum was separated by centrifugation at 1500 rpm for 15 minutes. The serum was frozen and kept at -20°C until hormonal assays could be carried out.

Kits of hormones used for human research:

Human PROG, PGE2, and AREG ELISA kits of (Sunlung Biotech. Co., LTD) products allow for the in vitro quantitative determination of human PROG(pg/ml), PGE2 (ng/ml), and AR(ng/ml) concentrations in serum and follicular fluid.

Materials provided:

Reagents include an Assay plate, Standards solution specific for each hormone, HRP-conjugate, Antibody specific for each hormone, Wash buffer, Substrate A, Substrate B, and Stop solution.

The procedure of (progesterone, prostaglandin E2, and Amphiregulin) was followed according to the manufacturer's recommendations.

Statistical analysis:

Collection, summarization, and analysis of data were done and presented using three statistical software programs: System -SAS (2012), Microsoft Office Excel 2016, and Duncan test (1955) polynomial. While categorical variables were presented as numbers and percentages based on the results of the Kolmogorov-Smirnov test of normalcy distribution for numerical variables, numerical variables were reported as mean and standard deviation (SD) or median and interquartile range (IQR). The association between categorical variables was examined using the Chi-square test, and any necessary adjustments were performed.

RESULTS:

Description of the age of the study groups enrolled in the present study:

Fifty subfertile women were involved and diagnosed to have polycystic ovary syndrome (PCOS) and 50 subfertile or infertile women who had no PCOS) male factor), there was no significant(P>0.05) difference in the mean age between the non-PCOS group and the PCOS group, (30.26±2.94versus 30.50±2.50years, P=0.6612), as shown in table (1).

The performance of the Kolmogorov-Smirnov normality test revealed that the age variable in the current study was not normally distributed. Hence the study estimated the median and interquartile range (IQR) of both groups and the entire sample for a more precise description of the age variable and the results are shown in Table (1).

Table 1: Mean age and age range in subfertile women classified into PCOS and non-PCOS groups

Characteristic	Non-PCOS (n = 50)	PCOS (n = 50)	P-value
Mean age ±SD (years)	30.26±2.94^a	30.50±2.50^a	0.6612 (N.S.)
Age range (years)	25-35	25-35
Median age (IQR) (years)	30	30

PCOS: polycystic ovary syndrome; n: number of cases; SD: standard deviation; IQR: interquartile range; NS: not significant.

The relationship of BMI with pregnancy rate:

In the PCOS group, the mean BMI for pregnant women was (27.850.94) and for non-pregnant women was (28.310.96), as shown in Table (2). In this study, the mean BMI for pregnant women was 27.231.13, and for non-pregnant women was 27.481.09. In the non-PCOS group, The pregnancy rate and the women's BMI did not differ statistically significantly.

Table 2: Relation of BMI with a pregnancy rate

	pregnancy	No.	BMI Mean± SD	significant
PCOS (n = 50)	positive	20	27.85±0.94^a	N.S.
PCOS (n = 50)	negative	30	28.31±0.96^a	N.S.
Non-pcos (n = 50)	positive	18	27.23±1.13^a	N.S.
Non-pcos (n = 50)	negative	32	27.48±1.09^a	N.S.

PCOS: polycystic ovary syndrome; n: number of cases; SD: standard deviation; NS: not significant.

Types of infertility:

Women were classified into those having primary infertility and those having secondary infertility and the results are shown in Table (3). The proportion of women with primary infertility was somewhat lower in the non-PCOS group than in the PCOS group, with 28 (percentage 45.2) against 34 (percentage 54.8).

On the other hand, the number and percentage of women complaining of secondary infertility were slightly more in the non-PCOS group than in the PCOS group, 22 (%57.9 )versus 16 (% 42.1), and there was a significant statical difference (P<0.05) in the mean of primary infertility of non- PCOS and PCOS group (28 (%45.2) versus 34 (%54.8), P=0.4461) and there was no significant(P>0.05) difference in the mean of secondary infertility of non-PCOS groups and PCOS group (22 (%57.9) versus 16 (%42.1), P=0.3304).

Biochemical pregnancy outcome in PCOS and non-PCOS groups:

Positive biochemical pregnancy was detected in 38 (38%) of all women participating in the current study. As shown in table (4). The percentage of pregnancy was higher in PCOS than in non-PCOS groups, (20 (%52.6) versus 18 (%47.4) However, there was no statical significant difference between positive and negative pregnancy in PCOS and non-PCOS groups.

Table 4: Percentage of pregnancy in PCOS and non-PCOS groups

Pregnancy

Non-PCOS

(n = 50)

PCOS

(n = 50)

Total

χ²

Positive

18 (%47.4)

20 (%52.6)

38 (%100)

0.105

n.s.

0.7456

Negative

32 (%51.6)

30 (%48.4)

62 (%100)

0.064

n.s.

0.7995

PCOS: polycystic ovary syndrome; n: number of cases; χ²: Chi-square statistic value; %: percentage

Comparison between PCOS patients and control in level of follicular and serum hormones and growth factors:

Highly significant differences were encountered between PCOS and non-PCOS groups regarding serum progesterone, prostaglandin E2 hormones at the day of ova pickup (OPU), follicular Progesterone, prostaglandin E2 hormones at the day of ova pickup (OPU), Amphiregulin growth factor at the day of ova pickup (OPU), progesterone, prostaglandin E2 hormones at the day of embryo transfer, Amphiregulin growth factor at the day of embryo transfer (table 5).

Progesterone (P4):

The mean of serum P4 levels on the day of ova pick up ± SE in both non-PCOS and -PCOS groups were(1.43±0.51 and0.99±0.28). While the mean of serum p4± SE in both non-PCOS and PCOS groups on the day of embryo transfer were (1.50±0.51 and 0.96±0.16), both P4 levels on the day of OPU and ET were significantly higher P(P<0.001) in non –PCOS group. The mean of FF P4 in non-PCOS and PCOS (1.87±0.56 and 1.55±0.83) was significantly higher than P (0.0260) in the non-PCOS group

Prostaglandin (PGE2):

On the day of ova pick-up, the mean serum PGE2 levels in the PCOS and non-PCOS groups were 9.97±2.60 and 7.04±1.55, respectively. There was a very significant (P0.0037) difference between the non-PCOS and PCOS groups, even though their respective mean serum PGE2± SE values on the day of ET were 9.97±2.60 and 8.81±2.23. FF PGE2 in non-PCOS and PCOS groups (6.92±0.73 and 9.55±1.91) was significantly higher(P<0.001) in PCOS than in the non-PCOS group (Figure 2).

Amphiregulin Growth factor (AREG):

The mean of serum AREG levels at the day of ova pick up ± SE in both non-PCOS and PCOS groups were (1.84±0.47and 1.39±0.38) respectively. The mean of serum AREG ± SE in both non-PCOS and PCOS groups on the day of embryo transfer was (1.47±0.19and 1.28±0.20) respectively. The levels of AREG in (FF, OPU, ET) were highly significant (P<0.001) in the non-PCOS group compared with the PCOS group.

Table 5: Comparison of hormones and growth factor between PCOS and non-PCOS groups on the day of (ET, OPU, and FF)

Hormone	Non-PCOS (n = 50)	PCOS (n = 50)	P*
Hormone	Mean± SD	Mean± SD	P*
S.PROG.OPU	1.43±0.51^a	0.99±0.28^b	<.0001 **
S.PGE2.OPU	7.56±1.88^a	5.47±1.63^b	<.0001 **
S.AREG.OPU	1.84±0.47^a	1.39±0.38^b	<.0001 **
S.PROG.ET	1.50±0.51^a	0.96±0.16^b	<.0001 **
S.PGE2.ET	4.98±2.82^a	3.49±2.63^b	0.0037 **
S.AREG.ET	1.47±0.19^a	1.28±0.20^b	<.0001 **
FF.PROG	1.87±0.56^a	1.55±0.83^b	0.0260 *
FF.PGE2	7.48±1.65^a	5.70±1.93^b	<.0001 **
FF.AREG	1.64±0.60^a	1.28±0.21^b	0.0001 **

Mann Whitney U test; n: number; SD: standard deviation; Prog : Progesterone: PGE2:Prostaglandin; CD2: cycle day 2; HS: highly significant; FF: follicular fluid; OPU: oocyte pickup; AREG: Amphiregulin growth factor.

Comparison Between the Pregnancy Outcome and Level of (Amphiregulin growth factor, Progesterone, and Prostaglandin) at the Day ova pick, and embryo transfer and in follicular fluid:

In the non-PCOS group, levels of AREG on the day of OPU, ET, and FF were significantly higher (P<0.001) in pregnant women. As shown in Figures (1 and 2), there was a highly significant increase in the level of AREG in pregnant more than in non-pregnant women in non- PCOS group, There was no statistically significant difference (P>0.05) between pregnant and non-pregnant women, with the exception of the AREG level in the PCOS group level on the day of ET, which was slightly higher in pregnant women than non-pregnant women.

Figure 1: Mean of Amphiregulin in (OPU, ET, and FF) between pregnant and non-pregnant women in the non-PCOS group

Figure 2: Mean of Amphiregulin in (OPU, ET, and FF) between pregnant and non-pregnant women in the PCOS group

In the non-PCOS group, there was no significant difference (P>0.05) in levels of P4 between pregnant and non-pregnant women on the day of OPU. The level of P4 on the day of ET was significantly higher (P<0.05) in pregnant more than in non-pregnant women. P4 FF level was highly significant (P<0.001) in pregnant women more than non-pregnant in the non-PCOS group. In the PCOS group, there was no significant difference (P>0.05) in levels of P4 on the day of OPU in serum and FF between pregnant and non-pregnant women. On the day of ET, the level of serum P4 was highly significant (P<0.001) in pregnant more than in non-pregnant women (Figures 3 and 4).

Figure 3: Mean of Progesterone in (OPU, ET, and FF) between pregnant and non-pregnant women in the non-PCOS group

Figure 4: Mean of Progesterone in (OPU, ET, and FF) between pregnant and non-pregnant women in the PCOS group

In PCOS and non-PCOS groups, there is a high statical significant difference (P<0.001) between pregnant and non-pregnant women in levels of PGE2 in FF, day of OUP, and ET (Figures 5 and 6).

Figure 5: Mean of Prostaglandin in (OPU, ET, and FF) between pregnant and non-pregnant women in the non-PCOS group

Figure 6: Mean of Prostaglandin in (OPU, ET, and FF) between pregnant and non-pregnant women in the PCOS group

DISCUSSION:

Clinical Characteristics of the Infertile Female:

Age:

It is recognized by many studies that age is well known as one of the most important factors predicting IVF/ICSI outcome ^32,
33,. In addition, with the increase in maternal age, IVF outcomes become increasingly worse ³⁴. Some IVF centers even restricted the last age for IVF to 43 years old ³⁵. In the present study, the patient's age was comparable in PCOS patients and non-PCOS groups. There was no significant difference in mean age in both the non-PCOS and PCOS groups (30.26±2.94versus 30.50±2.50)

Body Mass Index:

Menstrual problems and anovulation have been identified as mechanisms by which body mass impacts reproduction. This is likely due to the changed secretion of LH, sex hormone binding globulin, ovarian and adrenal androgen, as well as altered insulin resistance. Other processes may be proposed, such as the uterus's reduced receptivity after embryo transfer, which is likely due to disrupted endometrial activity ⁽³⁶⁾. In the present study, BMI was comparable in PCOS patients and non-PCOS- group, also there were no significant differences in BMI among pregnant and non-pregnant patients in both the PCOS and non-PCOS groups.

Hind A. Beydoun et al. ³⁷ found that although PCOS and continuous BMI appear to influence some of the process metrics of IVF/ICSI success, they had no appreciable effect on the rate of pregnancy, miscarriage, or live birth or on the number of mature oocytes fertilized per mature oocyte retrieved or inseminated.

Pregnancy outcome:

When compared to the non-PCOS group, the pregnancy rate among PCOS patients in this research was significantly higher. This outcome is consistent with earlier findings by Esinler et al. ⁽³⁸⁾ who discovered that appropriate pregnancy rates in ovulatory PCO and PCOS patients were caused by a sufficient number of harvested oocytes, fertilized oocytes, and transplanted high-quality embryos. Others, however, discovered that although the clinical abortion rate for PCOS patients was greater than controls, there was no change in the pregnancy rate. The authors hypothesized that in PCOS patients, only cytoplasmic maturity was affected, not nuclear maturity ³⁹.

Type of infertility:

According to the present study's findings, primary infertility is more common than secondary infertility in both women with and without PCOS and in the complete sample of women. These findings are consistent with those of several studies that examined the prevalence of primary and secondary infertility, finding that the former is more common than the latter ⁴⁰.

Progesterone:

The present study showed that there is no significant difference in serum progesterone among the pregnant and non-pregnant groups in PCOS and non-PCOS patients on the day of OPU. These results agreed with results obtained by Yasser Ibrahim Orief et al ⁴¹who found that the serum concentration of progesterone was similar in conception and non-conception cycles.

P4 levels in the non-PCOS group are significantly elevated in FF and on the day of OPU, which is consistent with the findings of LU Xiu-e et al. ⁴². Numerous authors discovered that oocyte maturation, subsequent implantation, and pregnancy were all predicted by a high concentration of progesterone in follicular fluid. ⁽⁴³⁾ A lack of progesterone during the luteal phase is linked to early miscarriages. Progesterone also regulates follicular rupture and luteinization, two processes that are essential for maintaining a healthy pregnancy throughout the early embryonic stage. Progesterone levels rise following the LH surge. Therefore, to ensure a good pregnancy, progesterone production needs to be tightly regulated ⁴⁴

Amphiregulin Growth Factor:

Our study shows that there was a highly significant increase in AREG levels in PCOS and non-PCOS groups of pregnant and non-pregnant women.

EGF, transforming growth factor-alpha (TGFa), amphiregulin (AREG), epiregulin, betacellulin, epigene, neuregulins, and heparin-binding EGF-like growth factor are all members of the family of receptor ligands for epidermal growth factor (EGF) ⁴⁵. EGF, TGFa, amphiregulin (AREG), epiregulin, betacellulin, epigene, neuregulins, and heparin-binding EGF-like growth factors are all examples of growth factors among the proteins that make up epidermal growth factor receptor ligand ⁴⁶ and Juengel JL³⁷ and by Downs Growth-factor family members have drawn a lot of interest lately due to their function in regulating ovulation and egg maturation ^{45, 47}.

Epidermal growth factor (EGF) receptor ligands include epidermal growth factor (EGF), transforming growth factor-alpha (TGFa), amphiregulin (AREG), epiregulin, betacellulin, epigene, neuregulins, and heparin-binding EGF-like growth factor.

It was recently demonstrated that AREG mediates the expression of steroidogenic acute regulatory protein (StAR) and progesterone production in human granulosa cells when hCG is present. Furthermore, there was a positive correlation between the quantity of AREG in the follicular fluid and the increase in serum progesterone between the day of OPU and the day following ET. These results demonstrate that progesterone levels, which are essential for a healthy pregnancy, may be predicted by AREG expression in the follicular fluid ^48–50.

Prostaglandin E2:

The results of the present study showed that serum PGE2 and FF PGE2 on the day of ova pickup were all significantly higher in women with positive pregnancy outcomes than those with negative pregnancy outcomes in both PCOS and non-PCOS groups.

A crucial part of mammalian reproduction, ovulation is a strictly controlled process. The LH surge controls a number of genes linked to the ovulatory response. Prostaglandin E2 (PGE2) is one of these ovulation-related genes that has been identified as a crucial mediator that controls ovulation. The limiting step in the generation of prostaglandins is catalyzed by the enzyme cyclooxygenase (COX). The granulosa cells of periovulatory follicles showed a considerable rise in COX-2 mRNA and protein levels following the treatment of human chorionic gonadotropin (HCG). Furthermore, following hCG injection, PGE2 concentrations in the follicular fluid rose. Our research shows that in human granulosa cells, AREG is more effective in promoting COX-2 expression and PGE2 synthesis ⁵¹.

The addition of luteinizing hormone (LH) stimulates the conversion of arachidonic acid to PGE2 in granulosa cells, and perfusion of the ovaries with PGE2 causes mature follicle rupture. The vascular permeability alterations that occur at the implantation site are thought to be mediated by prostaglandins. Prostaglandin levels have been found to rise dramatically near the implantation site⁵².

CONCLUSION:

This study found that with COX2 expression and PGE2 generation in primary human granulosa cells, AREG mediates the hCG-induced increase in progesterone production.

REFERENCES:

1. Practice Committee of the American Society for Reproductive Medicine. Definitions of infertility and recurrent pregnancy loss. Fertil Steril. 2008; 90: 560.

2. Jain G, Khatuja R, Juneja A, Mehta S. Laparoscopy: As a First Line Diagnostic Tool for Infertility Evaluation. Journal of Clinical and Diagnostic Research. 2014; 8(10): OC01-OC02.

3. Bhattacharya S, Johnson N, Tijani HA, Hart R, Pandey S, Gibreel AF. Female infertility. British Medical Journal. 2010; 10: 8-12.

4. Radhakrishnan SA. Polycystic Ovarian Syndrome (PCOS) in Adolescence. Asian J. Nur. Edu. & Research. 2012; 2(2): 55-64.

5. Sheela Williams, Lissa J, Saraswathi. K. N. A Study to Assess the Effectiveness of Structured Teaching Programme on Knowledge Regarding Polycystic Ovarian Disease among Adolescent Girls in Selected Colleges in Mysuru. Asian J. Nur. Edu. and Research. 2017; 7(3): 371-375.

6. Ashlesha Nagare, Mercy Anjore. To Assess the Effectiveness of Planned Teaching Programme on Knowledge regarding Polycystic Ovarian Syndrome, its Management and Prevention among Late Adolescent Girls in selected Undergraduate Colleges of Nagpur City. Asian J. Nursing Education and Research. 2019; 9(1): 01-03.

7. Nancychandra Priya P., Shwetha M. N.. Knowledge regarding Polycystic Ovarian Syndrome among Young Female Adults. Asian J. Nursing Education and Research. 2019; 9(1): 84-86.

8. Yashoda Shrivastava, Parvati Jagdev. A Study to assess the Effectiveness of self Instructional module on Knowledge regarding Polycystic Ovarian Syndrome among B.Sc. Nursing students of Selected nursing college. Asian J. Nursing Education and Research. 2019; 9(3): 388-390.

9. Sanasam Birjeni Devi, C. Susila. Diagnosis, causal factors and associated diseases of PCOS: A Mini-review. Asian Journal of Nursing Education and Research. 2022; 12(1): 138-3.

10. Usama Shoukath, Umama Shoukath, Salma Sultana, Mohammed Nayeem Uddin. Polycystic Ovary Syndrome (PCOS): A Thief of Womenhood. Asian J. Res. Pharm. Sci. 2018; 8(2): 68-72.

11. Akshay Gade, Rutuja Sawant, Shreya Parkar, Prajakta Kegade. Polycystic ovary syndrome: An Overview, Diagnosis and Treatment of PCOS. Asian J. Pharm. Tech. 2020; 10(4): 265-272.

12. Khushboo Brar, Tarundeep Kaur, P. Vadivukarrasi Ramanadin. Knowledge regarding Poly Cystic Ovarian Syndrome (PCOS) among the Teenage Girls. Int. J. Nur. Edu. and Research. 2016; 4(2): 136-140.

13. Asha K Varughese, Veena Greetta Tauro. Polycystic Ovarian Syndrome – An Overview. Int. J. Nur. Edu. and Research. 2019; 7(4): 601-604.

14. Aghajanova L, Hamilton A, Giudice L. Uterine Receptivity to Human Embryonic Implantation: Histology, Biomarkers, and Transcriptomics. Seminars in Cell and Developmental Biology. 2008; 19(2): 204-211.

15. Timeva T, Shterev A, Kyurkchiev S. Recurrent Implantation Failure: The Role of the Endometrium. Journal of Reproduction and Infertility. 2014; 15(4): 173-183.

16. Lim HJ, Dey SK. HB-EGF: a unique mediator of embryo-uterine interactions during implantation. Experimental Cell Research. 2009; 315(4): 619-626.

17. Su R-W, Fazleabas AT. Implantation and Establishment of Pregnancy in Human and Nonhuman Primates. Advances in Anatomy, Embryology and Cell Biology. 2015; 216: 189-213.

18. Liu L, Wang Y, Yu Q. The PI3K/Akt signaling pathway exerts effects on the implantation of mouse embryos by regulating the expression of RhoA. International Journal of Molecular Medicine. (2014); 33(5): 1089-1096.

19. Dekel N, Gnainsky Y, Granot I, Mor G. Inflammation and Implantation. American Journal of Reproductive Immunology. 2010; 63(1): 17-21.

20. Hantak AM, Bagchi IC, Bagchi MK. Role of Uterine Stromal-Epithelial Crosstalk in Embryo Implantation. The International Journal of Developmental Biology. 2014; 58(0): 139-146.

21. Godinjak Z, Bilalovic N. Estrogen and Progesterone Receptors in Endometrium in Women with Unexplained Infertility. Materia Socio-Medica. 2014; 26(1): 51-52.

22. Huang, Y. et al. Altered amphiregulin expression induced by diverse luteinizing hormone receptor reactivity in granulosa cells affects IVF outcomes. Reproductive Biomedicine Online 30, 593–601, doi: 10.1016/j.rbmo.2015.03.001 (2015).

23. Allen VM, Wilson RD, Cheung A.Pregnancy outcomes after assisted reproductive technology. J Obst Gynaecol Can. 2006; 28(3): 220-250.

24. 14-Borini A, Lagalla C, Cattoli M, et al. Predictive factors for embryo implantation potential. Reprod Biomed Online. 2005; 10: 653-68.

25. Dennis SJ,Thomas MA,Williams DB, et al. Embryo morphology score on day 3 is predictive of implantation and live birth rate. J Assist Reprod Genet. 2006; 23: 171-17.

26. Rimon-Dahari N, Yerushalmi-Heinemann L, Alyagor L, Dekel N.Ovarian folliculogenesis. Results and Problems in Cell Differentiation. 2016; 58: 167–190.

27. Verlhac MH. Meiosis and oogenesis. Mol Biol Cell. 2012; 23(6): 971.

28. Shimasaki S, Moore RK, Otsuka F and Erickson GF. The bone morphogenetic protein system in mammalian reproduction. Endocrine Review. 2004; 25: 72-10.

29. Muttukrishna S. and Ledger W. Inhibin, activin,and follistatin in human reproductive physiology. Endocrine,autocrine and paracrine actions of inhibin,activin and follistatin on follicle stimulating hormone. Imperial.College press, London, UK: 2001; 3:72.

30. Rodgers RJ, and Irving-Rodgers HF. Formation of the ovarian follicular antrum and follicular fluid.Biol Reprod. 2010; 82(6): 1021-1029.

31. Leroy JL, Vanholder T, Delanghe JR,et al.Metabolic changes in follicular fluid of the dominant follicle in high-yielding dairy cows early post partum.Theriogenology. 2004; 62: 1131-1143.

32. Van Loendersloot LL, van Wely M, Limpens J, et al. Predictive factors in in vitro fertilization (IVF): a systematic review and meta-analysis. Hum Reprod Update. 2010; 16: 577 –89.

33. Van Loendersloot LL, Repping S, Bossuyt PM, et al. Prediction models in in vitro fertilization; where are we? A mini review. Journal of Advanced Research. 2014; 5: 295–301.

34. Yan JH, Wu KL, Tang R, et al. Effect of maternal age on the outcomes of in vitro fertilization and embryo transfer(IVF-ET). Sci China Life Sci. 2012; 55: 694–8.

35. Hourvitz A, Machtinger R, Maman E, et al. Assisted reproduction in women over 40 years of age: how old is too old? Reprod Biomed Online. 2009; 19(4): 599-603.

36. Sathya A, Balasubramanyam S, Gupta S, et al. Effect of body mass index on in vitro fertilization outcomes in women. J Hum Reprod Sci. 2010; 3(3): 135-8.

37. Hind A. Beyound, Laurel Stadtmaur et al.Poly cystic Ovary Syndrome, Body Mass Index and Outcome of Assisted Reproductive Technologies. Reprod Biomed online. 2009; 18(6): 856-863.

38. Esinler,U.Bayar,G.Bozdag, and H.Yrali.Outcome of intracytoplasmic sperm injection in patients with polycystic ovary syndrome or isolated polycystic ovaries. Fertility and sterility. 2005; 84(4): 932-937.

39. Baoli Yin,Haoying Hao, Duo Wie et al.Patients with Polycystic Ovary Syndrome have Successful Embryo arrest. Int J Clin Exp Med. 2015; 8(4): 6247-6251.

40. Al-Turki H A.Prevalence of primary and secondary infertility from tertiary center in eastern Saudi Arabia. Middle East Fertility Society Journal. 2015; 20 (4): 237–240.

41. Yasser Ibrahim Orief, Tarek Abd Ezaher Karkor,Hisham Aly Saleh et al. Comparison between steroid expression in serum and follicular fluid in polycystic ovary patients and unexplained infertility patients undergoing assisted reproductive techniques. Middle East Fertility Society Journal. 2014; 19(1): 57-63.

42. LU Xiu-e, HUANG He-feng,LI Mei-gen, et al. Progesterone levels of serum and follicular fluid in non-obese patients with polycystic ovary syndrome during IVF cycles. J Zhejiang Univ SCI. 2005; 6B(9): 897-902.

43. Revelli A, Piane L, Casano S, et al. Follicular fluid content and oocyte quality: from single biochemical markers to metabolomics. Reprod Biol Endocrinol. 2009; 7: 40.

44. Ke, R. W. Endocrine basis for recurrent pregnancy loss. Obstetrics and gynecology clinics of North America 2014; 41, 103–112. doi: 10.1016/j.ogc. 2013; 10.003.

45. Ben-Ami I, Freimann S, Armon L, Dantes A, Ron-El R, Amsterdam A. Novel function of ovarian growth factors: combined studies by DNA microarray, biochemical and physiological approaches. Mol Hum Reprod. 2006; 12: 413–9.

46. Park JY, Su YQ, Ariga M, Law E, Jin SLC, Conti M. EGF-like growth factors as mediators of LH action in the ovulatory follicle. Science. 2004; 303:682.

47. Juengel JL, McNatty KP. The role of proteins of the transforming growth factor-beta superfamily in the intraovarian regulation of follicular development. Hum Reprod Update. 2005; 11: 144–61.

48. Zamah, A. M. et al. Human oocyte maturation is dependent on LH-stimulated accumulation of the epidermal growth factor-like growth factor, amphiregulin. Human Reproduction. 2010; 25, 2569–2578. doi: 10.1093/humrep/deq212

49. Huang, Y. et al. Altered amphiregulin expression induced by diverse luteinizing hormone receptor reactivity in granulosa cells affects IVF outcomes. Reproductive biomedicine online 30, 593–601, doi: 10.1016/j.rbmo.2015.03.001 (2015).

50. Ben-Ami, I. et al. EGF-like growth factors as LH mediators in the human corpus luteum. Human Reproduction 2009; 24, 176–184, doi: 10.1093/humrep/den359.

51. Fang, L., Cheng, J. C., Chang, H. M., Sun, Y. P. and Leung, P. C. EGF-like growth factors induce COX-2-derived PGE2 production through ERK1/2 in human granulosa cells. The Journal of Clinical Endocrinology and Metabolism. 2013; 98; 4932–4941. doi: 10.1210/ jc.2013-2662

52. Hester KE, Harper MJ, Duffy DM. Oral administration of the cyclooxygenase-2 (COX-2) inhibitor meloxicam blocks ovulation in non-human primates when administered to simulate emergency contraception. Hum Reprod. 2010; 25: 360 –367.

Received on 29.08.2022 Revised on 22.07.2024

Accepted on 30.06.2025 Published on 13.01.2026

Available online from January 17, 2026

Research J. Pharmacy and Technology. 2026;19(1):382-389.

DOI: 10.52711/0974-360X.2026.00056

This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Creative Commons License.

Infertility type	Non-PCO (n = 50)	PCO (n = 50)	Total (n =100)	χ²	P
Primary, n (%)	28 (% 45.2)	34 (% 54.8)	62(%100)	0.58 n.s.	0.4461
Secondary, n (%)	22 (%57.9 )	16 (% 42.1)	38 (%100)	0.94 n.s.	0.3304