Overview

Endometrium is the mucus membrane that lines the inside of the uterus. During ovulatory cycles, the endometrium undergoes cyclic changes and prepares for implantation. If implantation fails to occur, the endometrium sheds as a part of the menstrual cycle. A large body of data suggests that an efficient embryo-uterine “cross-talk” is critical for initiation and progression of successful implantation in humans.^1,2   This cross talk is recognized by a synchronous development and interaction between a hatched blastocyst and endometrium which serves to orchestrate implantation-related events during both natural and assisted conception cycles.³

Normally, the endometrium is non-receptive for an embryo. However, it becomes receptive for a particular period of time—called the implantation window—due to the influence of steroid hormones and paracrine signals from the developing embryo. While there is no consensus regarding the dates and duration of this implantation window, endometrial receptivity is believed to be optimal between cycle days 20 and 24, after which the endometrium becomes refractory to the implantation process. Investigators have identified several morphological and biochemical factors that influence endometrial receptivity during the implantation window (figure 1).^4,8   Derangements in the regulation of these factors may lead to endometrial anomalies that may jeopardize the success of implantation and pregnancy. In light of these data, the endometrium is considered a major fertility-determining factor.⁹

Endometrial thickness: A clinical perspective

Among the factors affecting implantation and conception, endometrial thickness is deemed critically important.¹⁰   It is utilized as an indicator of endometrial receptivity and serves as a valuable prognostic parameter for successful pregnancy in assisted reproduction cycles such as in vitro fertilization (IVF).¹¹

A myriad of investigators believe that an endometrial thickness less than a specific value is capable of reducing the likelihood of pregnancy to almost zero.¹²   However, the optimal endometrial thickness has remained an area of controversy, and no conclusive cut-off value has been established till date.¹¹   Some authors have suggested that a thickness of greater than 9 mm is associated with a more favorable prognosis for pregnancy in IVF while others have reported high conception rates above a cut-off value of 10 mm.^1,13   To further shed light on the optimal endometrial thickness during assisted reproduction, Aydin et al¹⁴   undertook a prospective cross-sectional study on 593 women undergoing IVF. The patients were divided into four groups according to endometrial thickness (ranging from <7 to >14mm) and implantation and pregnancy rates were evaluated. Although no threshold was observed above which pregnancy was unlikely to occur, both implantation and clinical conception rates were found to be remarkably lower in those with endometrial thickness <7 mm (figure 2). Similar results were obtained in a systematic review and meta-analysis¹⁵   which showed that the probability of clinical pregnancy for an endometrial line ≤7 mm was significantly lower compared to one greater than 7 mm (23.3% versus 48.1%) (figure 3).

The above-mentioned evidences make it reasonable to suggest that an endometrial thickness of <7mm is suboptimal for embryo transfer and may be associated with reduced pregnancy rates.¹⁶

Thin endometrium in IVF cycles

In the recent years, thin endometrium has captured attention due to its negative impact on pregnancy outcomes in IVF cycles.¹⁷   Accumulating evidences suggest that a thin endometrium reduces the chances of conception in relatively young patients (<38 years old), in patients requiring more than 10 days of gonadotropin stimulation, and in those with poor quality embryos undergoing embryo transfer cycles.¹⁸

An inadequate endometrium is characterized by abnormal epithelial growth, decreased vascular endothelial growth factor (VEGF) expression, and poor vascular development.¹⁹   Several lines of evidence suggest that blood flow in the endometrium is an important determinant of endometrial receptivity and implantation.²⁰   In an endometrial line of <7 mm, the functional layer is either very thin or absent, due to which the implanting embryo is much closer to the spiral arteries and the higher oxygen concentrations of the basal endometrium. This high oxygen tension near the basal layer is detrimental unlike the usual low oxygen tension of the surface endometrium, and may lead to implantation failure.²¹

Causes of thin endometrium

An enlarging body of evidence suggests that endometrial growth depends on factors such as a woman’s age, infertility etiology, drug protocol, serum estradiol level and previous injuries to endometrium. In some older women, myomatous uterus or pelvic inflammatory disease (PID) can affect IVF results. While age is an indirect factor affecting the endometrium, ovarian anomalies and estrogen levels have direct effects. Reduced endometrial thickness in IVF cycles can also be attributed to the presence of intrauterine adhesions and past surgical curettage procedures that can cause damage to the endometrium, leading to poor IVF outcomes. Besides, as previously discussed, poor blood supply to the uterine layers also signals an increased risk of implantation failure. A great deal of evidence suggests that uterine fibroids, arterial constriction of the uterus, and uterine fibroid embolization significantly contribute to compromised vascularity, subsequently causing reduced endometrial thickness. Figure 4 presents a composite depiction of the factors responsible for a thin endometrium.^22-25

Endometrial receptivity

Since endometrial thickness is an important biomarker of endometrial receptivity during the implantation window, it is logical to suggest that a thin endometrium would lead to compromised endometrial receptivity and consequently reduced implantation rates. The average rate of successful implantation in IVF programs is estimated to be nearly 25%. Pooled data from published trials suggests that poor endometrial receptivity is responsible for approximately two-thirds of implantation failures, while the embryo quality itself is responsible for only one-third of such cases.^8,26     Furthermore, in women with unexplained implantation failure, despite optimal endocrine response and endometrial development, good-quality embryos, and absence of a clinical pathology, compromised endometrial receptivity appears to be a major factor in inhibiting embryo attachment.²⁷

Effect of age on endometrial receptivity

Researchers have reported abnormal endometrial receptivity in ageing women. While there has been no conclusive evidence of age-induced histological changes in the endometrium, some authors have noted a significant fall in ongoing pregnancy rates in ARTs, from 48.8% in women aged <30 years to 13.6% in those aged <42 years. Likewise, the rate of successful implantation rate has been observed to drop from 29% at <34 years of age, to approximately 5% at age 42. Data derived from several other studies concerning ovum donation also point to a drop in conception rates with increasing recipient age. Albeit oocyte senescence is a key factor responsible for such a trend, there is convincing evidence that uterine factors such as demised endometrial receptivity play a significant role.²⁸

Hormonal requirement for endometrial receptivity and implantation

Appreciation of the importance of endometrial receptivity in ART has fuelled interest in understanding the role of ovarian steroidal hormones in boosting success outcomes. Investigators have accumulated a large body of data showing estrogen and progesterone to play pivotal roles in endometrial receptivity, blastocyst implantation, and sustenance of pregnancy. It is also apparent from assisted conception studies that the ratio of progesterone to estrogen in serum is important for the secretory maturation of the endometrial glands during the midluteal phase. These findings have been corroborated by both experimental and human studies.

Clinical reports delineating the effects of these hormones report that they function to mobilize several molecular modulators in a spatiotemporal manner thereby facilitating embryo attachment. Further, they regulate the cascade of growth factors and cytokines which are the prime paracrine mediators of embryoendometrial dialogue. Progesterone during the luteal phase, particularly orchestrates the synthesis and secretion of numerous proteins. diagnostic evaluation of endometrium during assisted conception.²⁹

Diagnostic evaluation of endometrial status

Although much has been learned about the factors regulating embryo-endometrial interactions, the ideal method to predict uterine receptivity is yet to be established. Histological evaluation of an endometrial biopsy specimen has remained the mainstay for assessing endometrial status. However, its invasiveness has been a major clinical limitation in context of ART cycles. Researchers suggest that estimation of estradiol and progesterone levels in circulation is non-invasive and has high utility in reflecting endometrial status. It is believed that the effect of serum estradiol on the endometrium is dictated by the number and function of endometrial estrogen receptors.³⁰   Investigators have observed a significant correlation between the thickness of the endometrium and the serum estradiol level in the follicular phase of natural ovulatory cycles; this finding reflects the diagnostic value of serum estradiol measurement in IVF programs also.³¹   Dickey et al³²   have further shown that serum estradiol levels are predictive of, not only the increasing maturity of endometrial pattern, but also the number of mature eggs and quality embryos. The functions of estrogen during the follicular phase, which include proliferation of surface epithelium, glands, stroma and endometrial vessels, and upregulation of progesterone receptors, sufficiently explain this finding.³³

Despite its broad utility, hormonal milieu assessment may sometimes be inaccurate. This has led to the development of more precise, non-invasive methods of receptivity assessment such as ultrasonography. Ultrasonographic endometrial thickness measurement, endometrial pattern investigation, endometrial volume computation, uterine and subendometrial blood flow analysis by Doppler sonography are few of the methods that can be used to evaluate uterine receptivity and consequently better predict pregnancy outcomes during ART programs. Over the last few years, transvaginal sonography has emerged as an important tool, greatly improving the ability of clinicians to appropriately diagnose endometrial defects.^30,34,35         Mortakis and colleagues³⁶   have reported transvaginal ultrasound to be an excellent initial diagnostic method with high sensitivity in detecting endometrial defects. The technique is rarely nondiagnostic and has added advantages of being painless and devoid of complications.³⁷

Management strategies

As previously mentioned, a thin endometrium adversely affects reproductive performance, due to which its appropriate management is crucial for women contemplating IVF. In many cases of infertility where there is no identifiable pathology, treatment to improve the poorly prepared endometrium has proved successful. However, achieving the optimal endometrial thickness remains a challenge in clinical practice.¹⁷

Several strategies have been undertaken to circumvent thin endometrium with the objective of increasing conception rates. There are three major types of interventions for thin endometrium; these include:

• Optimizing serum estradiol levels, with oral/ vaginal estradiol, low-dose human menopausal gonadotropin (HMG) or extended estrogen treatment
• Boosting uterine vascularity and correcting fibrosis, with low-dose aspirin, vaginal sildenafil citrate, pentoxifylline, vitamin E, l-arginine, electroacupuncture or neuromuscular electrical stimulation
• Employing novel approaches based on the use of bone marrow stem/progenitor cells or cytokine such as Granulocyte-Colony Stimulating Factor (G-CSF) for intrauterine administration.³⁸

Estrogen and progesterone

Among the above-mentioned strategies, artificial endometrial preparation, comprising induction of endometrial proliferation with estrogens and secretory transformation with progesterone, has gained importance.38 Investigators posit that estrogen and progesterone are necessary and sufficient for endometrial development towards a receptive milieu in ART settings such as cryopreserved embryo transfer cycles.³⁹

Estradiol valerate is an estrogen formulation, which has been available since the 1970s in oral and parenteral forms.⁴⁰   It is efficacious for the management of thin endometrium and is also superior to other estrogens (such as conjugated equine estrogens-CEE) in conferring greater benefit without significant safety concerns. It is considered to be less potent in terms of inducing hepatic protein synthesis and is thereby believed to induce fewer estrogen-related adverse drug effects.^41,42  >   The potential of estradiol valerate preparation in improving endometrial thickness has been evidenced by several researchers. A study⁴³   was conducted to ascertain the effect of its administration on endometrial lines during in vitro maturation cycles. The results revealed that treatment with estradiol valerate since the beginning of the menstrual cycle promoted an increase in endometrial thickness and volume both on the day of planned egg retrieval and one week later. Taking this notion a step further, Li and coworkers⁴⁴   demonstrated that treatment with oral estradiol valerate significantly improves ultrasonographic thickness of endometrium and enhances endometrial response by producing adequate plasma levels of estradiol without any adverse effects.

It is worth mentioning here that the micronized form of estradiol in particular offers several benefits (figure 5).^45,46      The Martindale drug reference⁴⁷   mentions micronized preparation of estradiol to be the only natural unconjugated estrogen which is orally active. Researchers have demonstrated oral estradiol valerate supplementation significantly thickens endometrial lining and additionally results in increased number of follicles and a high percentage of mature oocytes.⁴⁸

Vaginal route of estradiol administration is generally reserved for cases in which the oral route is not effective. The first-pass hepatic metabolism can be bypassed by utilizing vaginal route of estradiol administration. Vaginal use of micronized estradiol leads to significantly greater free serum estradiol levels when compared to those achieved after oral administration.^49,50      However, the pharmacodynamics of estradiol absorption via this route may not be satisfactory and researchers have observed attenuated absorption.⁵¹

Progesterone therapy also exerts a favorable impact on pregnancy rates in IVF by optimizing endometrial thickness and enhancing receptivity for implantation.⁵²   In IVF-embryo transfer cycles with egg donation, sequential administration of physiologic doses of estradiol and progesterone to women with poor ovarian function successfully restores endometrial receptivity and facilitates the establishment of viable pregnancy.²⁸   Similar results were reported in another study which showed that hormonal treatment combining estradiol and progesterone improves endometrial receptivity and ultimately yields higher pregnancy rates.⁵³

Human chorionic gonadotropin

A large body of data suggests that the human chorionic gonadotropin (hCG) receptor is present in endometrium and therefore a favorable interaction could be expected when hCG is administered in the proliferative phase of endometrial growth. In a proof of concept study, recipients of fresh donor or frozen embryos who had a consistently thin endometrium non-responsive to previous hormone treatments were enrolled. On day-8 or 9 of the estrogen administration, and continuing 8 mg estrogen per day, subcutaneous injections of 150 IU HCG were initiated daily for 7 days. After a week on HCG priming (day-14 or 15), endometrial thickness was monitored by ultrasound, and progesterone was initiated. The results revealed that majority of the patients (52.9 %) experienced more than 10 % improvement after hCG priming while 35.3 % had more than 20 % improvement (figure 6). It was concluded that 150 IU HCG endometrial priming for 7 days in the proliferative phase of estrogen substituted cycles for frozen embryos is highly promising, as not only the thickness of the endometrium improves but subsequently the receptivity also becomes near normal. However, large-scale studies are required to confirm these findings.⁵⁴

If adequate endometrium is not attained despite treatment with the above-mentioned regimens, it is worth evaluating the uterine cavity to rule out pathology.²⁸    Vaginal sildenafil may be tried empirically, preferably in conjunction with estradiol valerate. Sildenafil citrate, a type 5-specific phosphodiesterase inhibitor, induces vasodilatory effects, thereby improving uterine blood flow and endometrial status. Studies have demonstrated the combination of sildenafil and estradiol valerate to optimize endometrial thickness in 100% patients undergoing IVF cycles, with successful pregnancy in 75% of them.⁵⁵

Granulocyte colony stimulating factors

There is evidence derived from several studies that G-CSF stimulates neutrophilic granulocyte proliferation and differentiation, acts on macrophages of decidual cells, thereby influencing implantation.⁵⁶   Uterine perfusion with G-CSF represents an advanced treatment option for thin endometrium. Investigators have reported successful endometrial expansion to at least a minimal thickness of 7 mm after administration of G-CSF in patients refractory to treatment with other formulations.⁵⁷   To take this notion a step further, a randomized multicentric double blinded study⁵⁸   was undertaken on women undergoing IVF or ICSI. The cultured embryo was transferred to test media containing either G-CSF or control media. The results showed that the survival rate of the embryo cultured with G-CSF till 12th week and birth was significantly higher than controls. These data validate the efficacy of the G-CSF in ARTs.

Conclusion

Embryo implantation is the result of a well-orchestrated sequence of events which requires a receptive endometrium, a good-quality embryo at the blastocyst developmental stage and a synchronized embryo-maternal dialogue. It is rather surprising that endometrium is refractory to embryo attachment during most days of the menstrual cycle and becomes receptive only for a limited period, which is called the implantation window.

Endometrial thickness is utilized as an indicator of endometrial receptivity during the implantation window and is deemed critically important for the success of implantation and conception. Studies have shown suboptimal endometrial lines to be responsible for a significant proportion of pregnancy failures in ART cycles. Hence, over the recent years, strategies to optimize endometrial thickness in women contemplating IVF have gained momentum. The main treatment for thin endometrium include firstly, increasing serum estradiol levels, with oral estrogen preferably micronized estradiol valerate administration. Vaginal route may be tried for cases in which the oral route is not effective. If an adequate endometrium cannot be achieved with exogenous steroids, it is worth evaluating the uterine cavity to rule out pathology. If no pathologic abnormality is found, higher-dose estrogen for a longer period may be necessary. If an appropriate thickness still cannot be achieved other options like vaginal sildenafil preferably in conjunction with estradiol valerate, G-CSF may be tried.

References

1. Kovacs P, Matyas S, Boda K, Kaali SG. The effect of endometrial thickness on IVF/ICSI outcome. Hum Reprod. 2003 Nov;18(11):2337-41.
2. Paria BC, Song H, Dey SK. Implantation: molecular basis of embryo-uterine dialogue. Int J Dev Biol. 2001;45(3):597-605.
3. Timeva T, Shterev A, Kyurkchiev S. Recurrent Implantation Failure: The Role of the Endometrium. Journal of Reproduction & Infertility. 2014;15(4):173-183.
4. Sharkey AM, Smith SK. The endometrium as a cause of implantation failure. Best Pract Res Clin Obstet Gynaecol. 2003 Apr;17(2):289-307.
5. Harper MJ. The implantation window. Baillieres Clin Obstet Gynaecol. 1992 Jun;6(2):351-71.
6. Sharma A, Kumar P. Understanding implantation window, a crucial phenomenon. Journal of Human Reproductive Sciences. 2012;5(1):2-6.
7. Tabibzadeh S. Molecular control of the implantation window. Hum Reprod Update. 1998 Sep-Oct;4(5):465-71.
8. Firouzabadi DR, Davar R, Hojjat F, Mahdavi M. Effect of sildenafil citrate on endometrial preparation and outcome of frozen-thawed embryo transfer cycles: a randomized clinical trial. Iranian Journal of Reproductive Medicine. 2013;11(2):151-158.
9. Strowitzki T, Germeyer A, Popovici R, von Wolff M. The human endometrium as a fertility-determining factor. Hum Reprod Update. 2006 Sep-Oct;12(5):617-30.
10. Singh N, Bahadur A, Mittal S, Malhotra N, Bhatt A. Predictive value of endometrial thickness, pattern and sub-endometrial blood flows on the day of hCG by 2D doppler in in-vitro fertilization cycles: A prospective clinical study from a tertiary care unit. Journal of Human Reproductive Sciences. 2011;4(1):29-33.
11. Al-Ghamdi A, Coskun S, Al-Hassan S, Al-Rejjal R, Awartani K. The correlation between endometrial thickness and outcome of in vitro fertilization and embryo transfer (IVF-ET) outcome. Reproductive biology and endocrinology : RB&E. 2008;6:37.
12. Wang L, Qiao J, Li R, Zhen X, Liu Z. Role of endometrial blood flow assessment with color Doppler energy in predicting pregnancy outcome of IVF-ET cycles. Reproductive Biology and Endocrinology : RB&E. 2010;8:122.
13. Noyes N, Liu HC, Sultan K, Schattman G, Rosenwaks Z. Endometrial thickness appears to be a significant factor in embryo implantation in in-vitro fertilization. Hum Reprod. 1995 Apr;10(4):919-22.
14. Aydin T, Kara M, Nurettin T. Relationship between Endometrial Thickness and In Vitro Fertilization-Intracytoplasmic Sperm Injection Outcome. International Journal of Fertility & Sterility. 2013;7(1):29-34.
15. Kasius A, Smit JG, Torrance HL, Eijkemans MJ, Mol BW4, Opmeer BC, Broekmans FJ. Endometrial thickness and pregnancy rates after IVF: a systematic review and meta-analysis. Hum Reprod Update. 2014 Jul-Aug;20(4):530-41.
16. Eftekhar M, Sayadi M, Arabjahvani F. Transvaginal perfusion of G-CSF for infertile women with thin endometrium in frozen ET program: A non-randomized clinical trial. Iranian Journal of Reproductive Medicine. 2014;12(10):661-666.
17. Lebovitz O, Orvieto R. Treating patients with “thin” endometrium – an ongoing challenge. Gynecol Endocrinol. 2014 Jun;30(6):409-14.
18. Zhang X, Chen CH, Confino E, Barnes R, Milad M, Kazer RR. Increased endometrial thickness is associated with improved treatment outcome for selected patients undergoing in vitro fertilization-embryo transfer. Fertil Steril. 2005 Feb;83(2):336-40.
19. Miwa I, Tamura H, Takasaki A, Yamagata Y, Shimamura K, Sugino N. Pathophysiologic features of “thin” endometrium. Fertil Steril. 2009 Apr;91(4):998-1004.
20. Yilmaz N, Kiliç S, Madendağ Y, Madendağ I, Ozgün A, Ozakşit G, Batioğlu S. Endometrial parameters in IVF and IUI administration on elderly women. Turk J Med Sci. 2010; 40 (3): 343-348.
21. Zhao J, Zhang Q, Li Y. The effect of endometrial thickness and pattern measured by ultrasonography on pregnancy outcomes during IVF-ET cycles. Reproductive Biology and Endocrinology : RB&E. 2012;10:100.22.
22. Krzymowski T, Stefanczyk-Krzymowska S. Uterine blood supply as a main factor involved in the regulation of the estrous cycle–a new theory. Reprod Biol. 2002 Jul;2(2):93-114.
23. Ng EH, Chan CC, Tang OS, et al. Endometrial and subendometrial blood flow measured by three-dimensional power Doppler ultrasound in patients with small intramural uterine fibroids during IVF treatment. Hum Reprod. 2005;20(2):501-6.
24. Simms-Stewart D, Fletcher H. Counselling patients with uterine fibroids: a review of the management and complications. Obstet Gynecol Int. 2012;2012:539365.
25. Habibzadeh V, Nematolahi Mahani SN, Kamyab H. The correlation of factors affecting the endometrial thickness with pregnancy outcome in the IUI cycles. Iran J Reprod Med. 2011 Winter;9(1):41-6.
26. Achache H, Revel A. Endometrial receptivity markers, the journey to successful embryo implantation. Hum Reprod Update. 2006 Nov- Dec;12(6):731-46.
27. Fatemi HM, Popovic-Todorovic B. Implantation in assisted reproduction: a look at endometrial receptivity. Reproductive BioMedicine Online. 2013;27:530– 538.
28. Elnashar AM, Aboul-Enein GI. Endometrial receptivity. Middle East Fertility Society Journal. 2004;9(1):10-24.
29. Rashid, N. A., Lalitkumar, S., Lalitkumar, P. G. and Gemzell- Danielsson, K. Endometrial Receptivity and Human Embryo Implantation. American Journal of Reproductive Immunology. 2011 Jul;66:23–30.
30. Friedler S, Schenker JG, Herman A, Lewin A. The role of ultrasonography in the evaluation of endometrial receptivity following assisted reproductive treatments: a critical review. Hum Reprod Update. 1996 Jul-Aug;2(4):323-35.
31. Katayama T. Ultrasonographic changes in the endometrium during ovulatory cycles–correlation to serum estradiol and progesterone concentrations. Nihon Sanka Fujinka Gakkai Zasshi. 1990 Nov;42(11):1530-6.
32. Dickey RP, Olar TT, Curole DN, Taylor SN, Rye PH. Endometrial pattern and thickness associated with pregnancy outcome after assisted reproduction technologies. Hum Reprod. 1992 Mar;7(3): 418-21.
33. Narvekar SA, Gupta N, Shetty N, Kottur A, Srinivas M, Rao KA. The degree of serum estradiol decline in early and midluteal phase had no adverse effect on IVF/ICSI outcome. Journal of Human Reproductive Sciences. 2010;3(1):25-30.
34. Senturk LM, Erel CT. Thin endometrium in assisted reproductive technology. Curr Opin Obstet Gynecol. 2008 Jun;20(3):221-8.
35. Leone FP, Timmerman D, Bourne T, Valentin L, Epstein E, Goldstein SR, Marret H, Parsons AK, Gull B, Istre O, Sepulveda W, Ferrazzi E, Van den Bosch T. Terms, definitions and measurements to describe the sonographic features of the endometrium and intrauterine lesions: a consensus opinion from the International Endometrial Tumor Analysis (IETA) group. Ultrasound Obstet Gynecol. 2010 Jan;35(1):103-12.
36. Mortakis AE, Mavrelos K. Transvaginal ultrasonography and hysteroscopy in the diagnosis of endometrial abnormalities. J Am Assoc Gynecol Laparosc. 1997 Aug;4(4):449-52.
37. Davidson KG, Dubinsky TJ. Ultrasonographic evaluation of the endometrium in postmenopausal vaginal bleeding. Radiol Clin North Am. 2003 Jul;41(4):769-80.
38. Chen X, Chen S. Successful Pregnancy in Recurrent Thin Endometrium with New Uses for an Old Drug.
39. Burks H, Paulson R. Cryopreserved embryo transfer: endometrial preparation and timing. Semin Reprod Med. 2015 Mar;33(2): 145-52.
40. Micks E, Jensen JT. Estradiol valerate and dienogest: a novel fourphasic oral contraceptive pill effective for pregnancy prevention and treatment of heavy menstrual bleeding. Womens Health (Lond Engl). 2011 Sep;7(5):513-24.
41. Bissonnette F, Lussier-Cacan S, Fugère P, Bérubé S. Metabolic effect of two hormonal preparations in postmenopausal women. Maturitas. 1997 Jul;27(3):275-84.42.
42. Ndefo UA, Mosely N. Estradiol Valerate and Estradiol Valerate/ Dienogest (Natazia) Tablets: The First Four-Phasic Oral Contraceptive. Pharmacy and Therapeutics. 2010;35(11):614-617.
43. Martins W de P, Reis RM dos, Ferriani RA, de Araújo CHM, Nastri CO, Filho FM. Endometrial preparation for in vitro oocyte maturation: Early use of estrogen increases endometrial tissue and requires lower daily dosage: A cross over trial in “mock” cycles. Journal of Assisted Reproduction and Genetics. 2006;23(5):241-246.
44. Li TC, Cooke ID, Warren MA, Goolamallee M, Graham RA, Aplin JD. Endometrial responses in artificial cycles: a prospective study comparing four different oestrogen dosages. Br J Obstet Gynaecol. 1992 Sep;99(9):751-6.
45. Peter HM. Long term adherence to continuous combined HRT. Seven-year update on the Heikkinen study. Menopause international. 2003;9:8-9.
46. ‘Timing is everything, say new guidelines’ ppt. Data on record.
47. Sweetman SC, editor (2008) Martindale: The Complete Drug Reference, London, Pharmaceutical Press, Electronic version, (Edition 2008).
48. Elizur SE, Son WY, Yap R, Gidoni Y, Levin D, Demirtas E, Tan SL. Comparison of low-dose human menopausal gonadotropin and micronized 17beta-estradiol supplementation in in vitro maturation cycles with thin endometrial lining. Fertil Steril. 2009 Sep;92(3): 907-12.
49. Lete I, Dueñas JL, Esplugues JV, Marti-Cabrera M. Is the vagina an adequate route for the administration of hormonal contraceptives? Curr Drug Metab. 2010 Dec;11(10):839-49.
50. Tourgeman DE, Slater CC, Stanczyk FZ, Paulson RJ. Endocrine and clinical effects of micronized estradiol administered vaginally or orally. Fertil Steril. 2001 Jan;75(1):200-2.51.
51. Devroey P, Pados G. Preparation of endometrium for egg donation. Human Reproduction Update. 1998;4(6):856–861.
52. Segal S, Casper RF. Progesterone supplementation increases luteal phase endometrial thickness and oestradiol levels in in-vitro fertilization. Hum Reprod. 1992 Oct;7(9):1210-3.
53. Klkind-Hirsch KE, Phillips K, Bello SM, McNicho M, de Ziegler D. Sequential hormonal supplementation with vaginal estradiol and progesterone gel corrects the effect of clomiphene on the endometrium in oligo-ovulatory women. Hum Reprod. 2002 Feb;17(2):295-8.
54. Papanikolaou EG, Kyrou D, Zervakakou G, Paggou E, Humaidan P. “Follicular HCG endometrium priming for IVF patients experiencing resisting thin endometrium. A proof of concept study.” Journal of Assisted Reproduction and Genetics. 2013;30(10):1341-1345.
55. Sher G, Fisch JD. Vaginal sildenafil (Viagra): a preliminary report of a novel method to improve uterine artery blood flow and endometrial development in patients undergoing IVF. Hum Reprod. 2000 Apr;15(4):806-9.
56. Kunicki M, Łukaszuk K, Woclawek-Potocka I, Liss J, Kulwikowska P, Szczyptanska J. Evaluation of Granulocyte Colony-Stimulating Factor Effects on Treatment-Resistant Thin Endometrium in Women Undergoing In Vitro Fertilization. BioMed Research International. 2014;2014:913235.
57. Gleicher N, Vidali A, Barad DH. Successful treatment of unresponsive thin endometrium. Fertility and Sterility. 2011 May;95(6):2123.e13–2123.e17.
58. Kumar P, Mahajan S. Preimplantation and postimplantation therapy for the treatment of reproductive failure. Journal of Human Reproductive Sciences. 2013;6(2):88-92.