News & Events

27 Oct 2019


Digenic etiology of Müllerian anomalies in the female reproductive system


Cell Research published the work — Joint utilization of genetic analysis and semi-cloning technology reveals a digenic etiology of Müllerian anomalies on October 18, and the authors proposed and proved that the synergistic effect of “GEN1+WNT9B” digenic mutations plays an important part in Müllerian anomalies (MA).

The research was led by Professor HUA Keqin from the Obstetrics and Gynecology Hospital affiliated to Fudan University, in collaboration with Professor ZHANG Feng’s team, also from the Obstetrics and Gynecology Hospital and Professor LI Jinsong’s team from Shanghai Institute of Biochemistry and Cell Biology of Chinese Academy of Sciences. 

To nip MA in the bud

MA, a complex congenital disease, refers to a wide variety of anomalies in the female reproductive system: uterus, cervix, vagina, etc., and it usually comes with urinary system anomalies. Prof. Hua points out that clinical manifestations of MA include different types of anomalies in the reproductive system, such as no uterus or vagina, unicornuate uterus, rudimentary uterine horn and vaginal atresia. Vaginal anomalies alone happen to one in every 4,500 girls, and that is a conservative estimate based on patients recorded. MA has a negative impact on the physical and mental health of teenage girls and cast a shadow over their life in the long run.

For nearly a decade, Prof. Hua’s team has conducted minimally-invasive reconstructive surgeries on patients with female reproductive tract anomalies. They’ve conducted the first laparoscopic surgery of unicornuate uterus with rudimentary uterine horn in the world, enabling patients to bear their own kids. Considering the privacy, economic condition, and physical and mental needs of patients, they have even turned to biomesh and autogenous tissue to fix cervix and vagina malformations.

However, after a full recovery, patients can still be traumatized both physically and mentally from the disease. The team thought: Since MA is congenital, could it be possible to nip the disease in the bud?

Mutual inspiration and strong collaboration

Clinical practice drives basic research, which may, in return, inspires and even revolutionizes diagnosis and treatment once applied to practice. For years, researchers have been trying to find the pathogenesis of MA. Many genes have been spot to take part in the development of Müllerian ducts, some of which have been identified as candidate genes leading to MA.

Yet oligogenic disorders can hardly account for the human diseases that tend to be more complicated. There is an urgent need to find out whether the etiology of the disease is digenic or oligogenic. Therefore, Prof. Hua’s team have probed into MA in collaboration with Prof. Zhang’s and Prof. Li’s teams. Prof. Zhang has been committed to the basic and translational research on human genomic copy number variants (CNVs) and diseases caused by genetic variation such as infertility and birth defects. Prof. Li has succeeded in establishing mouse models that simulate human diseases, locating key genes that determine embryogenesis and key amino acids that determine the function of protein, through haploid embryonic stem cells and CRISPR-Cas9 genetic editing technology. The three teams forged a powerful alliance, burrowing into MA.

Unveiling the mystery of the etiology of MA

The research was based on phenotypes of 125 blood samples from MA patients collected by Prof. Hua’s team. Prof. Zhang’s team conducted targeting qPCR analysis and comparative genomic hybridization (CGH) microarray analysis on the samples. In total, 9 cases carrying potential pathogenic CNVs were identified. 

Digenic/ Oligogenic Models

According to Prof. Zhang, some patients’ unaffected mothers were found to carry only single deleterious variant either in GEN1 or WNT9B, which proved that a single variant was insufficient to cause MA. Researchers identified a frameshift mutation in WNT9B in the carrier of GEN1/2p24.2 deletion and a missense mutation in GATA3 in the carrier of TBX6/16p11.2 deletion, using whole exome sequencing (WES) in the 9 CNV carriers. Therefore, researchers speculated that “GEN1 + WNT9B” and “TBX6 + GATA3” might respectively suggest a potential digenic etiology of MA.

The speculation was further verified by genetically modified mouse models which simulated human diseases through semi-cloning technology and CRISPR-Cas9. What is worth noting is that it marks the world’s first MA research using semi-cloning technology and CRISPR-Cas9.

Experimental procedures for producing mouse models of double heterozygous mutants through semi-cloning and CRISPR-Cas9

Confirming that “GEN1 + WNT9B” could lead to MA in mouse models, they found, however that single genetic variant of either GEN1 or WNT9B was insufficient for MA manifestation in MA cohort and mouse models, while double heterozygous mutations in Gen1 and Wnt9b could lead to abnormal development of Müllerian ducts as a result of the synergistic effect caused by the two variants.

Their research provides important theoretical guidance for genetic counseling and molecular diagnosis. Accordingly, the team will screen other pathogenic variant combinations responsible for MA and better serve patients with genital malformations who want to have kids through assisted reproductive technology, such as embryo transfer. Their findings will provide more proof for the digenic etiology of MA and contribute to the “Healthy China” national strategy at the genetic level.


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