Unexplained recurrent pregnancy loss and unexplained infertility: twins in disguise

Is B-cell CLL/lymphoma 6 (BCL6) endometrial expression, a surrogate biomarker of endometriosis, elevated in women with unexplained recurrent pregnancy loss (uRPL) and unexplained infertility (UI) compared to fertile subjects?

Endometrial BCL6 expression is elevated to a similar degree in women with uRPL and UI compared to fertile controls.

Endometriosis has been linked to the genesis of endometrial progesterone resistance and to specific nuclear proteins, including endometrial BCL6. BCL6 overexpression (immune histologic score > 1.4) has been strongly associated with poor reproductive outcomes in IVF cycles in women with UI. Our previous data have demonstrated an accuracy of 94% for diagnosing endometriosis, and BCL6 protein is elevated in the decidua of women with uRPL.

In this case-control study, at a tertiary university teaching hospital, 110 samples (control n = 28; uRPL n = 29; UI n = 53) from pathological archives were analyzed. Timed endometrial biopsies were obtained between 2 January 2002 and 31 December 2016.

LH-timed endometrial biopsies were obtained from women with UI, uRPL (two or more consecutive losses) and normal fertile subjects during the mid-secretory phase of the menstrual cycle. Endometrial BCL6 protein levels were compared in women with UI and uRPL and fertile controls using western blot analysis and immunohistochemistry (HSCORE).

The mean age of the uRPL group was significantly higher than the others [mean (SD)] control = 32.7 (2.6); uRPL = 35.8 (3.7); UI = 32.7 (4.4); P = 0.002, ANOVA]. Seventy-nine percent of women in both subfertile groups (uRPL and UI, 65 out of 82) displayed elevated BCL6 protein levels. From these, a subset of cases with abnormal BCL6 went to laparoscopy and endometriosis was found in 9 out of 11 cases of uRPL and in 20 out of 21 cases of UI. Median BCL6 HSCORE for controls versus uRPL and UI was significantly different [median (interquartile); control = 0.3 (0.02 to 0.5); uRPL = 3 (1.9 to 3.6); UI = 2.9 (1.6 to 3.1); P < 0.0001, Kruskal-Wallis]. A significant trend in the association between the degree of infertility (fertile, uRPL and UI) and the HSCORE level (negative, medium and high) was found (P < 0.001; x ² for trend). Western blot of representative samples from each group demonstrated similar findings based on protein levels in the whole endometrium. After running ANCOVA analysis for age difference, the BCL6 difference among groups was still significant (P-value < 0.0001).

We studied subjects with two consecutive pregnancy losses rather than the definition adopted in Europe of three losses. The findings may lack external validity in other clinical settings (e.g. low prevalence of endometriosis).

Based on the data presented here, we postulate that the degree of BCL6 expression may represent a continuum of progesterone resistance and response to inflammation that occurs in women with endometriosis, yielding different degrees of infertility, from uRPL to UI.

This study was supported by NICHD/NIH R01 HD067721 (SLY and BAL), by Coordenação de Aperfeiçoamento de Pessoal de Nível Superior: Grant 99999.003035/2015-08 (BAL) and by CAPES/PROAP (RFS). Two authors (BAL, SLY) have licensed intellectual property for the detection of endometriosis. Dr Bruce Lessey is an unpaid scientific Advisor for CiceroDx. The other authors report no conflict of interest.