In 2016, Italian investigators reported that HHV-6A was detected in the uterus of approximately 40% of women with unexplained infertility while the virus was not found in any fertile women tested (Marci 2016). Now, a second study, led by Chicago fertility specialist Carolyn Coulam, MD, has confimed a potential role for HHV-6 in female infertility. In the present study, 37% of women experiencing recurrent implantation failure (RIF) were found to harbor HHV-6 in their endometrial tissue compared to 0% of fertile controls.
The team of investigators collected endometrial biopsies from 30 women who had undergone in vitro fertilization and failed to become pregnant after 4 or more embryo transfers. The biopsies were taken in a mock frozen embryo cycle after 5 days of progesterone treatment or on the expected day of frozen embryo transfer. Control tissue from women who had given birth at least once and had no history of infertility was collected 7-9 days after the luteinizing hormone surge. This collection time point corresponds to the luteal phase of the ovarian cycle and the secretory phase of the uterine cycle. Marci et al. reported that although HHV-6A DNA was present in HHV-6 positive tissues during all phases of the menstrual cycle, HHV-6A IE2 early protein was only expressed during the secretory phase.
Total RNA was assessed by quantitative RT-PCR, and HHV-6A/B U94 expression was measured. Expression of this viral transcript was confirmed in 37% of the women with RIF but in none of the fertile controls (p=0.038). The team also analyzed the endometrial immune profile by using qRT-PCR to quantify the expression of IL-18, IL-15, TNFSF12, TNFRSF12A, and CD56, and based on the expression patterns, the immune profile was designated as normal, overactivated, or low-activated. Among the women with implantation failure, the prevalence of abnormal endometrial immune profile types did not significantly differ between those with and without HHV-6. Altogether, 10/11 HHV-6+ RIF samples, 14/19 HHV-6- RIF samples, and only 1/10 control samples displayed abnormal endometrial immune profiles.
The group also measured expression levels of CD3e, CD19, CD16a, CD16b, and CD57 and found significantly higher levels of neutrophil-specific CD16b mRNA in HHV-6+ samples compared to HHV-6- samples. In addition, immunostaining of CD16b revealed an increased presence of neutrophils in the HHV-6 positive tissues. In contrast, expression of the NK and T-cell markers (CD3e, CD16a, CD56, and CD57) were not different between the populations. Transcription of the B-cell-related CD19 was significantly lower in HHV-6- RIF samples compared to control tissues, but while expression of CD19 also trended lower in HHV-6+ RIF samples compared to controls, the difference was not significant.
This independent reproduction of the findings of Marci et al. detailing a similar prevalence of HHV-6 in women who are unable to conceive (40% vs 37%), as well as an absence of HHV-6 in the endometrium of fertile controls (both 0%), is a compelling development in the effort to understand the roles of HHV-6A and –B in the female reproductive system. While the virus was not typed in this study, all of the HHV-6 in the former study was HHV-6A. Both studies also found variation in uterine immune markers between HHV-6+ and HHV-6- samples. Although the earlier results revealed an altered NK cell profile, with decreased CD56brightCD16- NK cells in HHV-6+ samples, and the present study did not find differences in total CD56 expression across HHV-6+ and negative samples, the authors note that this may be explained by the difference in methods, which did not allow for differentiation of NK cell subtypes. Further analysis of the uterine immune profile will help to clarify the ways in which HHV-6 is able to affect the endometrium, and on a larger scale, implantation and fetal development.
Looking forward, controlled clinical trials using an anti-HHV-6 agent to treat HHV-6+ women with unexplained infertility or RIF will be very useful in determining whether the virus is a causative factor behind female infertility. As noted by the authors, if HHV-6 is found to cause or contribute to RIF and/or unexplained infertility, treating the virus could improve the chance of positive pregnancy outcomes, and reduce the emotional and financial toll of infertility, in up to 40% of women with these conditions.
Dr. Coulam is in private fertility practice at the Reproductive Medicine Institute in Chicago. She collaborated with the clinical immunology laboratory at Rosalind Franklin University of Medicine and Science for the HHV-6 testing. Commercial HHV-6 testing for endometrial samples for HHV-6A/B DNA and mRNA is available at Coppe Labs in the US and at IKDT laboratories in Germany. Messenger RNA testing for HHV-6 is also available at the Clinical Immunology Laboratory of Rosalind Franklin University.
Find the full paper here: Coulam 2018.