Novel phenotypes and loci identified through clinical genomics approaches to pediatric cataract.

Pediatric cataract is highly heterogeneous clinically and etiologically. While mostly isolated, cataract can be part of many multisystem disorders, further complicating the diagnostic process. In this study, we applied genomic tools in the form of a multi-gene panel as well as whole-exome sequencing on unselected cohort of pediatric cataract (166 patients from 74 families). Mutations in previously reported cataract genes were identified in 58% for a total of 43 mutations, including 15 that are novel. GEMIN4 was independently mutated in families with a syndrome of cataract, global developmental delay with or without renal involvement. We also highlight a recognizable syndrome that resembles galactosemia (a fulminant infantile liver disease with cataract) caused by biallelic mutations in CYP51A1. A founder mutation in RIC1 (KIAA1432) was identified in patients with cataract, brain atrophy, microcephaly with or without cleft lip and palate. For non-syndromic pediatric cataract, we map a novel locus in a multiplex consanguineous family on 4p15.32 where exome sequencing revealed a homozygous truncating mutation in TAPT1. We report two further candidates that are biallelically inactivated each in a single cataract family: TAF1A (cataract with global developmental delay) and WDR87 (non-syndromic cataract). In addition to positional mapping data, we use iSyTE developmental lens expression and gene-network analysis to corroborate the proposed link between the novel candidate genes and cataract. Our study expands the phenotypic, allelic and locus heterogeneity of pediatric cataract. The high diagnostic yield of clinical genomics supports the adoption of this approach in this patient group.

Uterine cyclin A2 deficient mice as a model of female early pregnancy loss.

Proper action of the female sex steroids, 17ß-estradiol (E2) and progesterone (P4) on endometrium is essential for fertility. Beyond its role in regulating the cell cycle, cyclin A2 (CCNA2) also mediates E2 and P4 signaling in vitro, but a potential role in modulating steroid action for proper endometrial tissue development and function is unknown. To fill this gap in our knowledge, we examined human endometrial tissue from fertile and infertile women for CCNA2 expression and correlated this with pregnancy outcome. Functional assessment of CCNA2 was validated in vivo using a conditional Ccna2 uterine deficient mouse model while in vitro function was assessed using human cell culture models. We found that CCNA2 expression was significantly reduced in endometrial tissue, specifically the stromal cells, from women undergoing in vitro fertilization who failed to achieve pregnancy. Conditional deletion of Ccna2 from mouse uterine tissue resulted in an inability to achieve pregnancy which appears to be due to alterations in the process of decidualization, which was confirmed using in vitro models. From these studies, we conclude that CCNA2 expression during the proliferative/regenerative stage of the menstrual cycle acts as a safeguard allowing for proper steroid responsiveness, decidualization and pregnancy. When CCNA2 expression levels are insufficient there is impaired endometrial responsiveness, aberrant decidualization and loss of pregnancy.

Inflammatory MicroRNAs and the Pathophysiology of Endometriosis and Atherosclerosis: Common Pathways and Future Directions Towards Elucidating the Relationship.

Emerging data indicates an association between endometriosis and subclinical atherosclerosis, with women with endometriosis at a higher risk for cardiovascular disease later in life. Inflammation is proposed to play a central role in the pathophysiology of both diseases and elevated levels of systemic pro-inflammatory cytokines including macrophage migration inhibitory factor (MIF), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) are well documented. However, a thorough understanding on the mediators and mechanisms which contribute to altered cytokine expression in both diseases remain poorly understood. MicroRNAs (miRNAs) are important post-transcriptional regulators of inflammatory pathways and numerous studies have reported altered circulating levels of miRNAs in both endometriosis and atherosclerosis. Potential contribution of miRNA-mediated inflammatory cascades common to the pathophysiology of both diseases has not been evaluated but could offer insight into common pathways and early manifestation relevant to both diseases which may help understand cause and effect. In this review, we discuss and summarize differentially expressed inflammatory circulating miRNAs in endometriosis subjects, compare this profile to that of circulating levels associated with atherosclerosis when possible, and then discuss mechanistic studies focusing on these miRNAs in relevant cell, tissue, and animal models. We conclude by discussing the potential utility of targeting the relevant miRNAs in the MIF-IL-6-TNF-α pathway as therapeutic options and offer insight into future studies which will help us better understand not only the role of these miRNAs in the pathophysiology of both endometriosis and atherosclerosis but also commonality between both diseases.

Long non-coding RNAs in endometrial physiology and pathophysiology.

The endometrium is an essential component of the female uterus which provides the environment for pregnancy establishment and maintenance. Abnormalities of the endometrium not only lead to difficulties in establishing and maintaining pregnancy but also play a causative role in diseases of endometrial origin including endometriosis and endometrial cancer. Non-coding RNAs are proposed to play a role in regulating the genome in both normal endometrial physiology and pathophysiology. In this review, we first provide a general overview of non-coding RNAs and reproductive physiology of the endometrium. We then discuss the role on non-coding RNAs in normal endometrial physiology and pathophysiology of endometrial infertility. We then conclude with non-coding RNAs in the pathophysiology of endometriosis and endometrial cancer.

Increased CXCL12 expression in endometrium of women with abnormal uterine bleeding is post-transcriptionally mediated via miR-23b-3p and is associated with decreased expression of the miR-23b-3p/24-3p/27b-3p cluster: a pilot study.

OBJECTIVE: To study C-X-C motif chemokine 12 (CXCL12) and CXCR4 expression in endometrial tissue from both women with and without abnormal uterine bleeding (AUB) of endometrial origin and evaluate their relationship with microRNA (miRNA). DESIGN: Retrospective and laboratory study. SETTING: University-based research laboratory. PATIENT(S): Nine women with and without abnormal uterine bleeding, all of whom were in the secretory stage of their menstrual cycle, who provided endometrial biopsy tissue. INTERVENTION(S): Immunohistochemical localization of CXCL12 and CXCR4 as well as quantitative reverse-transcription polymerase chain reaction (qRT-PCR) assessment of mRNA expression in archived endometrial biopsy tissue and in vitro cell culture using the immortalized endometrial stromal cell line, t-HESC. Endometrial stromal cell line, t-HESC transfection with nontargeting, negative control miRNA mimics or miRNA mimics for miR-23b-3p and mRNA assessment miR-23b-3p expression confirmed by qRT-PCR and evaluation of impact on CXCL12 expression at the protein level by enzyme-linked immunosorbent assay and mRNA levels by qRT-PCR. MAIN OUTCOME MEASURE(S): Expression of CXCL12 and CXCR4 protein via immunohistochemistry and mRNA and miRNA levels of CXCL12 and CXCR4 as well as miR-23b-3p, miR-24b-3p, and miR-27b-3p, respectively, via qRT-PCR. RESULT(S): CXCL12 and its receptor CXCR4 expression were up-regulated in the endometrial tissue of women with AUB at the protein level, but this up-regulation of expression was only associated with increased CXCR4 mRNA expression. To evaluate whether CXCL12 may be post-transcriptionally regulated, we assessed expression of miR-23b-3p, a bona fide post-transcriptional regulator of CXCL12 expression. The expression of miR-23b-3p was statistically significantly lower in AUB endometrial tissue, as were fellow cluster members miR-24-3p and miR-27-3p. Transfection of t-HESC cells with pre-miR-23b-3p mimics statistically significantly reduced the levels of CXCL12 secreted protein but not mRNA levels, suggesting that miR-23b-3p retards protein translation independent of transcript degradation. CONCLUSION(S): Reduced expression of the miR-23b-3p/24-3p/27b-3p cluster is associated with elevated expression of CXCL12, which may contribute to the pathophysiology of AUB.