Middle East Fertility Society Journal Middle East Fertility Society ISSN: 1110-5690 Vol. 10, Num. 1, 2005, pp. 22-28

Middle East Fertility Society, Vol. 10, No. 1, 2005, pp. 22-28

OPINION

Characteristics and possible function of pinopodes seen on the surface of the receptive human endometrium

Anneli C. Stavreus-Evers, Ph.D.

Department of Clinical Science, Division of Obstetrics and Gynecology Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden
Address: Department of Clinical Science, Division of Ob. Gyn. K57, Karolinska University Hospital, SE-141 86, Stockholm, Sweden, Phone: +46 8 585 875 08, Fax: +46 8 585 875 75, E-mail: Anneli.Stavreus-Evers@klinvet.ki.se

Received on January 7, 2005;
revised and accepted on February 6, 2005

Code Number: mf05002

ABSTRACT

A synchronized development of embryo and endometrium is a prerequisite for a successful implantation. During the time of implantation, pinopodes appear on the endometrial surface. The precise function of these structural markers of endometrial receptivity is not known, but it is generally believed that they play a role in the implantation process. Further understanding of the function of these biomarkers and their role in embryo implantation could aid in better diagnosis and treatment of infertile couples in the future.

Keywords: Implantation window, pinopodes, endometrium.

INTRODUCTION

The endometrium is changing throughout the menstrual cycle in order to prepare for the implantation of an embryo. The maturation of the endometrium is dependent on interplay between the steroid hormones estrogen and progesterone. The endometrium is only receptive for blastocyst implantation for a short period of time, so called "implantation window" before and after which implantation does not occur (1).

Pinopodes, believed to be ultra structural markers of endometrial receptivity, appear on the endometrial surface at the time of implantation (Figure 1). Pinopodes was first observed in mice and rats (2,3) but similar structures have also been found in other species (4). Johannisson and Nelson were the first to report pinopode-like structures in human endometrium (5).

The morphological appearance of pinopodes differs between species. In rat and mouse, the pinopodes arise from the cell surface on stalks, which rise above the level of microvilli (6,7). In the rat, pinopodes are present in approximately 20% of the epithelial cells, whereas in human endometrium, pinopode structure seems to involve the majority of the non-ciliated epithelial cells (4,8).

Cellular organelles, such as mitochondria, have been found in pinopodes from several species (9,10) including human (11). Also considerable amount of glycogen is part of the content of pinopodes, which suggest that the formation of pinopodes is an energy consuming process (12).

Determination of the implantation window

Traditional dating according to the criteria of Noyes et al has been the standard method since the article was first published in 1950, and it still remains the most frequently cited article in the literature on infertility (13). The method is not perfect and detailed quantitative histological techniques have been used to study endometrial receptivity and function (14,15). However, none of these techniques has proven optimal for determination of endometrial receptivity and function.

Determination of the LH surge, measured in blood or urine, correlates well with histological dating according to Noyes et al (14). There has also been an attempt to date the endometrium by using integrins as biochemical markers (16). However the authors concluded that the morphological dating according to Noyes was superior.

The possibility to use pinopodes as markers of endometrial receptivity has been discussed from the early 1980s (17) and this was generally accepted in the mid 1990s (8,18). The embryo hatches on day 6 after the LH surge and is subsequently ready for implantation, probably within the nearest 24 hours. This corresponds to the formation of pinopodes in most women.

The length of the implantation window

The length of the implantation window is controversial. The implantation period is usually assumed to coincide with cycle day 20-22 in a standardized cycle of 28 days (19). However, subsequent evaluation of data based on highly sensitive hCG measurements in ART cycles estimated that the window of implantation lasts for approximately 5 days and extends from postovulatory days 6 to10 (cycle day 20-24 of an idealized 28 day cycle) (20).

The lifespan of pinopodes is also controversial. Our studies show that pinopodes are present in endometrium obtained on days LH+6 to LH+10 (21-24). 

In 60 biopsies obtained during the luteal phase, we found pinopodes from day LH+6 to LH+10 and they were most abundant on LH+7 (Figure 2). The duration of the pinopode appearance in each individual woman cannot be determined by obtaining only one biopsy in a cycle, but it can be speculated that the pinopodes appear on days LH+6 and/or LH+7 and disappear on days LH+8 and/or LH+9.

There is only one study, where sequential endometrial biopsies have been obtained from healthy during the same cycle. The number of women included in the study is small, only 10 women where the biopsies were been obtained in a normal cycle (25,26). This study suggests a lifespan of pinopodes less than 48 hours. However, there are possibilities that serial biopsies could cause changes in the endometrium such as a healing process or inflammation process that might influence the results.

There are some studies suggesting the presence of pinopodes from day LH+5 to LH+11(27) or that pinopodes exist throughout the luteal phase (28-31). However, none of these studies have used additional methods to verify their findings. Therefore, the morphological appearance of pinopodes of great importance and is currently debated. The morphology of the protrusions seen on the surface is of importance, since it might not be appropriate to regard any protrusion on the endometrial surface as a pinopode. However, the morphology of the protrusions varies during the late luteal phase (31) and other events, such as apoptosis, might be mistaken for pinopodes.

PINOPODES  

Hormonal regulation of pinopodes

In rats and mice, where endometrial receptivity requires both estrogen and progesterone, development of pinopodes is dependent on progesterone, and is inhibited by estrogen (32). In humans, the highest frequency of pinopodes is found in response to progesterone alone (32,33)

The pinopodes are present on the endometrial surface, when the serum levels of progesterone are high (21). It is possible to inhibit pinopode formation by administration of antiprogestins (34).

Increased serum progesterone concentration and down regulation of glandular progesterone receptor B expression are concurrent with development of pinopodes at the time of implantation (21).

Function of pinopodes

So far, the morphology of pinopodes has been described in detail (21, 25), but the biological function is less understood. The number of pinopodes varies between women, and a correlation between number of pinopodes and implantation rate has been demonstrated (25)

The function of pinopodes was first demonstrated in mice. An electron-dense tracer (ferritin) was introduced into the uterine lumen and the tracer was taken up into large vacuoles in the epithelial cells, which demonstrated the pinocytotic function of these protrusions (6). This fact named these structures pinopodes, which in Greek means "drinking foot". The pinocytotic process seen in rats and mice has not been shown in other species (10).

Uterine secretion is reduced during the time of implantation (35), but in contrast to mice and rats, pinopodes in human endometrium do not display pinocytotic function (36). This suggests that other mechanisms are involved in the reduction of uterine fluid e.g. less secretion or increased uptake.

The pinopodes arise form apical surface of the epithelial cells, and are large enough to trap the cilia. The pinopode structures might thereby prevent the blastocyst to be "swept away" by the cilia. This would facilitate close contact between the embryo and the endometrial surface.

It is possible that the embryo adheres to the pinopodes. In an in vitro systems blastocysts adhered to pinopodes on the surface (37). However, later in vivo studies by the same group also showed adhesion of the embryo to epithelial cells not forming pinopodes (38).

There are several biochemical markers related to the implantation window but very few markers have been related to the presence of pinopodes.

Integrins

Integrins are surface glycoproteins composed of α and β subunits that serve as receptors for components of extra cellular matrix such as osteopontin, fibronectin and collagens. These components have the capacity to act as bridging molecules between the blastocyst and the endometrial surface during the adhesion phase of the implantation process.  Integrins appear on the blastocyst as well as on the surface of glandular and luminal endometrial epithelium at the time of implantation (39). Fully developed pinopodes and strong intensity of alpha v beta 3 integrin is coinciding in endometrium obtained during the window of implantation (40).            

LIF and LIF receptor

Leukemia inhibitory factor plays an important role in the implantation process. In the mouse it has been shown that maternal endometrial LIF is an absolute pre-requisite for embryo implantation. (41). It has been shown that infertile women have less LIF in the uterine secretion than women with proven fertility (42). Increase of LIF in the glandular epithelium and upregulation of LIF receptor expression in the luminal epithelium has been demonstrated in the presence of pinopodes (24).  

HB-EGF

HB-EGF is a stimulator of cell proliferation, cell migration and cell motility (43). HB-EGF is highly expressed in the glandular and luminal epithelial cells at the time of implantation (23,44,45). HB-EGF is found both in the luminal epithelial cells of the endometrium and on the surface of the pinopodes (23). There are several possible functions of HB-EGF in the implantation process such as signaling between the endometrium and the blastocyst, to facilitate adhesion of the blastocyst to the luminal epithelial surface, and to heal the luminal epithelial surface after the invasion of the embryo.

Glutaredoxin

Glutaredoxin is expressed in the pinopodes, suggesting a role of these markers for the implantation process (22). The function is not known, but it can be speculated that these markers protect the endometrium and pre-implantation embryo from oxidative stress.

The importance of being able to determine when the endometrium is receptive is essential during IVF treatment. There are women, which do not become pregnant even after receiving a number of good quality embryos. For those women, with repeated implantation failure, there is reason to suspect a defect in endometrial development.

Two main types of defect endometrial receptivity have been described. The first is a delayed receptivity, accompanied by histological retardation (46,47). The second is the endometrium histologically in place but with a defect biochemical alteration (48).

The possibility to detect when the endometrium is receptive would be of great clinical value. However, there are still only few studies, where attempts to predict the endometrial receptivity have been performed. In one study, endometrial biopsies were obtained in one cycle before the embryo transfer (ET) and the ET time was estimated and according to the appearance of pinopodes. The study showed a possibility to predict the implantation window, and thereby increase the implantation rate (49)

Pinopodes are present on the endometrial surface at the time of implantation. The formation of pinopodes correlated to the increase in progesterone and decrease on PRB. However, further studies are needed to determine the function of pinopodes.

Further unravelling of molecules involved in the implantation process is needed for a better comprehension of the link between endometrial development and endometrial receptivity. Understanding the function of pinopodes and their role in implantation would result in more efficient diagnosis and treatment of infertile couples in the future.

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