Endometrial biopsy

The endometrial biopsy can be obtained without anesthesia and, if indicated, may be combined with a diagnostic hysteroscopy. The endometrial biopsy is then preserved in formalin. As only little tissue is removed, the risks (injury, infection, bleeding) are very low.

In our experience patients respond differently to the biopsy: some feel no discomfort, while others feel a brief discomfort similar to the one experienced during period. Buscopan® and / or Voltaren® given 1 hour before the biopsy may help to relief. After the biopsy, the tissue is sent directly to Mannheim in our laboratory in special shipping material provided by us.

Uterine natural killer cells

Uterine natural killer cells are immune cells that play an important role in both the second half of the cycle (luteal phase) and in early pregnancy. They are called "killer" due to their ability to target viruses and bacteria as well as tumor cells. By contributing about 70% of immune cells in the first trimester of pregnancy at the feto-maternal interface, the uNK cells represent the most significant population of all immune cells (2).

Results of international studies on "uterine natural killer cells and pregnancy" show conflicting results (3). Overall however, an increasing body of evidence suggests that an elevated number of natural uterine killer cells may have a negative impact on implantation or pregnancy (1,4,5).

After analyzing more than 20 000 endometrial biopsies and conducting scientific studies on uNK cells, approximately 20-30% of women with recurrent implantation failure (RIF) or recurrent miscarriage (RSA) show an increase of uNK cells (6).

Uterine plasma cells

Uterine plasma cells accumulate in the endometrium in case of chronic endometritis.

Women with chronic endometritis are usually asymptomatic or show mild or nonspecific symptoms such as chronic pelvic pain, pain during sexual intercourse, irregular vaginal bleeding, and persistent vaginal discharge. This explains why it remains undiagnosed in many cases (7).

The surface marker CD138 is used to identify uterine plasma cells. Chronic endometritis is diagnosed in international studies when only one CD138-positive plasma cell is found, as well as when many plasma cells are found (8-10). This also explains why the incidence of chronic endometritis in women with RSA or RIF ranges from 9 to 57.8% in prior research (8,11-17).

According to our analysis of over 20 000 endometrial biopsies with a focus on chronic endometritis, the incidence among women with RIF and RSA is approximately 10-15%.



Uterine regulatory T-cells

Regulatory T-cells are T-lymphocytes that play an important role in immune system self-regulation by preventing an excessive immunological response, for example, against the body's own cells.

Uterine regulatory T cells regulate the activity of the immune system and can promote tolerance towards the embryo. These, like uterine natural killer cells, play a vital immunological role during implantation and the early stages of pregnancy.

The importance of regulatory T cells in the context of implantation and early pregnancy has been examined in various studies (18-21). Patients with RSA and RIF were found to have significantly less uterine regulatory T-cells in the endometrium than healthy controls (22-24). This indicates that a decrease in uterine regulatory T-cells may impair uterine receptivity (also known as endometrial receptivity) and embryo implantation.

Endometrial BCL6 expression

Background: What is BCL6?

B-cell lymphoma 6 (BCL6) is a nuclear protein used to diagnose B-cell lymphoma. BCL6 not only has crucial functions in cell cycle control, cell differentiation and inhibition of apoptotic

processes (25-26), it also stimulates the expression of pro-inflammatory cytokines (27-29). Chronic inflammation can result in implantation and bleeding disorders, as well as dysmenorrhea. Endometriosis is linked to inflammatory processes, increased endometrial proliferation, decreased apoptosis and altered cellular immunity. The estrogen dominance that is often present is also associated with a reduced response to progesterone (so-called progesterone resistance) (30-32).

Recently, a large number of publications on BCL6 expression in women with fertility disorders and/or endometriosis patients have been published (33-43). They were able to demonstrate that endometrial BCL6 overexpression can be detected more frequently in patients with endometriosis and that it has a progesterone-antagonistic effect on the endometrium. BCL6 overexpression impairs the endometrium throughout the implantation process and is thus suggested as a biomarker for a dysfunctional endometrium and a critical causal component for progesterone resistance.

How to analyse endometrial BCL6 expression?

The level of BCL6 expression in the endometrial glands can be determined semi-quantitatively using immunohistochemistry. The percentage of stained endometrial gland nuclei and staining intensity are assessed and used to calculate the so-called HSCORE. A BCL6 expression with an HSCORE > 1.4 is pathologic /abnormal (33, 34,37-40). The endometrial biopsy should be performed 7-10 days after ovulation in a natural cycle.

BCL6 Expression and fertility

There are numerous studies that focus on BCL6 expression in fertility patients with/without endometriosis (33-40). These results show a high sensitivity (93%) and specificity (96%) for the presence of endometriosis (39). In a prospective cohort study, 75% of patients with unexplained infertility had BCL6 overexpression in the endometrium, which was associated with a significantly decreased pregnancy and live birth rate (17.3 vs. 64.7% and 11.5 vs. 58.8%, respectively) (34). Furthermore, patients with RIF and RSA showed a significantly higher expression of BCL6 in the endometrium compared to healthy control women (35,37).

Therapeutic options

Treatment in case of increased natural killer cells

To date, there are no approved immunological therapies targeting natural killer cells in patients with recurrent miscarriage or recurrent implantation failure. Different treatment options are currently under investigation in international studies. These options include glucocorticoids (e.g., prednisolone), lipid infusions (e.g., Intralipid®) and immunoglobulins.

Most studies on the treatment of elevated uterine killer cells use glucocorticoids and lipid infusions, but at present the therapies represent a so-called “off label” use (19-21).

Glucocorticoids

In a study with women suffering from idiopathic recurrent miscarriages and elevated uterine killer cells, administration of 20 mg prednisolone orally daily from cycle day 1-21 resulted in a significant reduction in uterine killer cells (19). Nevertheless, it should be kept in mind that glucocorticoids may also cause side effects, such as the development of gestational diabetes, arterial hypertension, preterm labor, decreased birth weight and disorders of pediatric neurological development and cleft lip and palate (22-24).

Lipid infusions

Lipid infusions contain soybean oil and have been used for many years in the treatment of intensive care patients for nutritional therapy. Studies have shown that lipid infusions may lower the activity of natural killer cells in peripheral blood (25-28). A study focusing the administration of lipid infusions in women with elevated uterine killer cells does not yet exist so far. No side effects have been reported for the use of lipid infusions in the present studies in patients with implantation failure or miscarriage. However, an allergy to soy, peanuts and egg yolks must be ruled out before administration of soy-containing lipid infusions.

Treatment in case of elevated plasma cells or detection of chronic endometritis

Chronic endometritis is treated with antibiotics. Following this treatment, a decrease in the inflammatory response has been shown and is linked to an increased live birth rate (7,8,12). Among others, the broad-spectrum antibiotic doxycycline is usually recommended for 14 as first line therapy (12). Different doses are used in the studies (e.g. 100 mg doxycycline twice daily for 14 days (8,29)). Alternative therapy regimens may include antibiotics such as azithromycin, metronidazole, ciprofloxacin, levofloxacin, or amoxicillin/clavulanic acid.

Treatment in case of decreased numbers of uterine regulatory T-cells

It was demonstrated that the intrauterine administration of HCG led to an increase in uterine Tregs. Studies suggest that the administration of HCG to patients with RIF who show a reduction in uterine Tregs might be a possible new therapeutic approach (30, 31).

The new studies confirm the data that:

• a subgroup of RM and RIF patients have significantly fewer Tregs in the endometrium, which may have a negative effect on endometrial receptivity
• the intrauterine administration of HCG represents a possible therapeutic option (individual approach, off-label use) in order to increase the recruitment of uterine Tregs.

A potential approach is based on the results of a meta-analysis, which included the data of 15 randomized controlled studies with 2,763 patients, and recommends the administration of 500 IU HCG within 15 minutes before embryo transfer (32).

Treatment options in case of BCL6 overexpression

Likes et al. assessed the effect of pharmacological (GnRH agonist suppression for 2 months) or surgical (laparoscopy) therapy prior to embryo transfer in women with unexplained infertility and BCL6 overexpression in a prospective cohort study (38). The live birth rate was significantly higher in the treatment groups than in women who remained untreated. Also, during the ASRM Meeting 2020, further interim results on medical and surgical treatment based on data from 7 centers and 189 patients were presented, supporting the previously published data (44).

Your doctor will discuss potential therapy options with you, taking into account your personal history and individual risk factors.



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