**1. Introduction**

52 Enhancing Success of Assisted Reproduction

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> The physiologic roles of both follicle stimulating hormone and luteinizing hormone are well established in the natural menstrual cycle. Research by Ryan and colleges in the 1960s established the concept of two different cells in the ovarian follicle, the thecal and granulosa cells, functioning in different manners to produce products of the steroid pathway, the "two cell hypothesis" (1, 2). Further work over the next two decades established the "two-cell two-gonadotropin" theory, demonstrating the action of FSH on granulosa cells and LH on thecal cells (3). Thecal cells alone were shown to express CYP17, the gene encoding for the critical enzyme in the conversion of progesterone and pregnenalone to androgens (3). Conversely, granulosa cells were demonstrated to be the cell expressing aromatase, allowing for the conversion of the androgens derived from the thecal cells to be converted to estrogens. The cooperation of both cells under the influence of both gonadotropins is essential for normal folliculogenesis and steroidogenesis in the ovary.

> LH has several physiologic roles within the ovary in addition to its roll in androgen production (Figure 1). LH receptor activation leads to increases in adenylate cyclase and cAMP, resulting in increased mitochondrial transport of cholesterol necessary for steroidogeneis through upregulation of StAR (4, 5). LH activity also induces the expression of EGF-like growth factors amphiregulin and epiregulin from luteinized granulosa cells (6). These factors protect these cells from apoptosis, induce pro-survival signaling cascades, and are critical in peri-ovulatory events (6, 7). The mid-cycle LH surge causes a cascade of events leading to ovulation of the oocyte from the ovarian follicle and take the oocyte out of meiotic arrest (8). Finally, LH receptors have been demonstrated in the endometrium during the implantation window, raising a possible roll for LH in peri-implantation endometrial events

© 2012 Hill and Propst, licensee InTech. This is an open access chapter distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2012 Hill and Propst, licensee InTech. This is a paper distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

(9, 10). The specific importance of LH activity can be demonstrated in patients with LHβ or LH receptor gene mutations. Case reports of these male and female patients have demonstrated hypogonadism, infertility, pseudohermaphroditism, and amenorrhea (11-13).

The Use of rLH, HMG and hCG in Controlled Ovarian Stimulation for Assisted Reproductive Technologies 55

have LH levels > 1 IU/L, a level presumably capable of driving adequate steroidogenesis (21). While the majority of patients have adequate endogenous LH levels to have successful ART cycles without exogenous LH, the value of additional exogenous LH administration has been a matter of debate. This chapter will review the scientific evidence surrounding the administration of exogenous LH in various forms (rLH, hMG, hCG) and its affect on ART

There are theoretical benefits of the use of exogenous LH for the oocyte and the endometrium. The putative purpose of controlled ovarian stimulation in ART is to maximize the number of oocytes retrieved. However, the evidence is clear that the addition of LH is not associated with an increase in the number of oocytes or the number of mature metaphase II oocytes (MII) retrieved. Indeed, the use of hMG has been shown to decrease the number of follicles, oocytes, and metaphase II oocytes (MII) as compared to rFSH alone (21, 22, 24-28), presumably due to the action of LH contained in hMG. This is confirmed by similar data comparing rLH plus rFSH versus rFSH alone which has shown a decrease in developing follicles and oocytes retrieved with rLH (29, 30). In the majority of these trials, the decrease was in oocytes from small to intermediate follicles, and the number of oocytes retrieved from large follicles and the number of MIIs retrieved were not different. This suggests the possibility that the use of exogenous LH activity is associated with a decreased in the development of small follicles which may have been unlikely to yield a fertilized 2PN.

In a series of *in vivo* studies evaluating the effect of LH activity on follicle growth, Filicori and colleges confirmed the findings that LH activity can decrease the growth of small follicles without impacting the continued growth and maturity of larger follicles. First, they demonstrated that the number of follicles under 10mm in size during ART stimulation positively correlated with FSH dose (r=0.193, p<0.05) but negatively correlated to LH dose (r=0.648, p<0.0001) (31). In another study, it was demonstrated that incrementally decreasing the dose of FSH from day 7 of stimulation and increasing the dose of LH resulted in a decrease in the number of follicles <10mm in size, without affecting follicles over 14mm in size (32). To evaluate if this effect was due to the decreasing FSH dose or the increasing LH dose, they performed a similar experiment where FSH was held steady at 150IU per day and patients were placed into groups of incrementally increasing LH doses. In this experiment, increasing doses of LH (in the presence of a constant dose of FSH) was again associated with a decrease in number of small follicles while not affecting the larger follicles (33). When the experiments were repeated utilizing hMG, hMG was also associated with a decrease in small follicles (34). These experiments and the results of many randomized controlled trials demonstrate that any beneficial effect of LH activity is not the result of an increase in oocyte

While the number of total oocytes, especially from small follicles, appears to be diminished in ART cycles utilizing LH, the quality of those oocytes may be increased. While direct

**2. Potential mechanisms of exogenous LH benefit in ART** 

There appears to be no negative effect on the development of larger follicles.

outcomes.

yield.

**Figure 1.** Key actions of LH within the ovary on the thecal cells, oocyte, and granulosa cells. Actions mediated by LH are indicated in red.

In assisted reproduction technologies (ART), the importance of LH is demonstrated clearly in hypogonadotropic hypogonadic patients. Patients with a profound lack of endogenous LH fail to undergo complete follicular maturation in the absence of exogenous LH (14, 15). Such patients require the exogenous administration of both LH and FSH to optimize reproductive outcomes (4, 16, 17). Urinary human menopausal gonadotropins were initially utilized in assisted reproductive technologies. These preparations were islolated and purified from large pools of human urine. One of the early urinary hMG products was Pergonal 75. One ampule of Pergonal 75 contained 75 international units (IU) of FSH and 75 IU of LH, which became an industry standard for ampules (18). These urinary hMG preparations contained both FSH and LH, as well as some hCG, and therefore patients were stimulated with both gonadotropins. Later advancements in monoclonal antibody technology enabled the production of urinary purified FSH and a more purified hMG, which is still used today (19, 20). Recombinant DNA technology using a mammalian cell culture system (Chinese hamster ovary cells) was used to produce recombinant human FSH, which was first licensed in 1995, and quickly replaced urinary FSH products. Recombinant human LH was later produced (18).

Despite the clear biologic importance of LH outlined in the preceding paragraphs, numerous studies have demonstrated successful ART outcomes with the use of exogenous FSH only (21, 22). A likely explanation is that LH is a very potent hormone, activating the LH receptor for adequate ovarian steroidogenesis when only 1% of LH receptors are bound (23). Even after GnRH agonist or antagonist down-regulation, a majority of patients will have LH levels > 1 IU/L, a level presumably capable of driving adequate steroidogenesis (21). While the majority of patients have adequate endogenous LH levels to have successful ART cycles without exogenous LH, the value of additional exogenous LH administration has been a matter of debate. This chapter will review the scientific evidence surrounding the administration of exogenous LH in various forms (rLH, hMG, hCG) and its affect on ART outcomes.
