Biallelic CPAMD8 variants are a frequent cause of childhood and juvenile open-angle glaucoma

Purpose: Developmental abnormalities of the ocular anterior segment in some cases can lead to ocular hypertension and glaucoma. CPAMD8 is a gene of unknown function recently associated with ocular anterior segment dysgenesis, myopia, and ectopia lentis. We sought to assess the contribution of biallelic CPAMD8 variants to childhood and juvenile open-angle glaucoma. Design: Retrospective, multicenter case series. Participants: A total of 268 probands and their relatives with a diagnosis of childhood or juvenile open-angle glaucoma. Methods: Patients underwent a comprehensive ophthalmic assessment, with DNA from patients and their relatives subjected to genome, exome, or capillary sequencing. CPAMD8 RNA expression analysis was performed on tissues dissected from cadaveric human eyes. Main outcome measures: Diagnostic yield within a cohort of childhood and juvenile open-angle glaucoma, prevalence and risk of ophthalmic phenotypes, and relative expression of CPAMD8 in the human eye. Results: We identified rare (allele frequency -5) biallelic CPAMD8 variants in 5.7% (5/88) of probands with childhood glaucoma and 2.1% (2/96) of probands with juvenile open-angle glaucoma. When including family members, we identified 11 individuals with biallelic variants in CPAMD8 from 7 unrelated families. Nine of these individuals were diagnosed with glaucoma (9/11, 81.8%), with a mean age at diagnosis of 9.22±14.89 years, and all individuals with glaucoma required 1 or more incisional procedures to control high intraocular pressure. Iris abnormalities were observed in 9 of 11 individuals, cataract was observed in 8 of 11 individuals (72.7%), and retinal detachment was observed in 3 of 11 individuals (27.3%). CPAMD8 expression was highest in neural crest-derived tissues of the adult anterior segment, suggesting that CPAMD8 variation may cause malformation or obstruction of key drainage structures. Conclusions: Biallelic CPAMD8 variation was associated with a highly heterogeneous phenotype and in our cohorts was the second most common inherited cause of childhood glaucoma after CYP1B1 and juvenile open-angle glaucoma after MYOC. CPAMD8 sequencing should be considered in the investigation of both childhood and juvenile open-angle glaucoma, particularly when associated with iris abnormalities, cataract, or retinal detachment

SVEP1 as a Genetic Modifier of TEK-Related Primary Congenital Glaucoma.

Purpose: Affecting children by age 3, primary congenital glaucoma (PCG) can cause debilitating vision loss by the developmental impairment of aqueous drainage resulting in high intraocular pressure (IOP), globe enlargement, and optic neuropathy. TEK haploinsufficiency accounts for 5% of PCG in diverse populations, with low penetrance explained by variable dysgenesis of Schlemm's canal (SC) in mice. We report eight families with TEK-related PCG, and provide evidence for SVEP1 as a disease modifier in family 8 with a higher penetrance and severity. Methods: Exome sequencing identified coding/splice site variants with an allele frequency less than 0.0001 (gnomAD). TEK variant effects were assayed in construct-transfected HEK293 cells via detection of autophosphorylated (active) TEK protein. An enucleated eye from an affected member of family 8 was examined via histology. SVEP1 expression in developing outflow tissues was detected by immunofluorescent staining of 7-day mouse anterior segments. SVEP1 stimulation of TEK expression in human umbilical vascular endothelial cells (HUVECs) was measured by TaqMan quantitative PCR. Results: Heterozygous TEK loss-of-function alleles were identified in eight PCG families, with parent-child disease transmission observed in two pedigrees. Family 8 exhibited greater disease penetrance and severity, histology revealed absence of SC in one eye, and SVEP1:p.R997C was identified in four of the five affected individuals. During SC development, SVEP1 is secreted by surrounding tissues. SVEP1:p.R997C abrogates stimulation of TEK expression by HUVECs. Conclusions: We provide further evidence for PCG caused by TEK haploinsufficiency, affirm autosomal dominant inheritance in two pedigrees, and propose SVEP1 as a modifier of TEK expression during SC development, affecting disease penetrance and severity

Glaucoma spectrum and age-related prevalence of individuals with FOXC1 and PITX2 variants

Published online 3 May 2017Variation in FOXC1 and PITX2 is associated with Axenfeld-Rieger syndrome, characterised by structural defects of the anterior chamber of the eye and a range of systemic features. Approximately half of all affected individuals will develop glaucoma, but the age at diagnosis and the phenotypic spectrum have not been well defined. As phenotypic heterogeneity is common, we aimed to delineate the age-related penetrance and the full phenotypic spectrum of glaucoma in FOXC1 or PITX2 carriers recruited through a national disease registry. All coding exons of FOXC1 and PITX2 were directly sequenced and multiplex ligation-dependent probe amplification was performed to detect copy number variation. The cohort included 53 individuals from 24 families with disease-associated FOXC1 or PITX2 variants, including one individual diagnosed with primary congenital glaucoma and five with primary open-angle glaucoma. The overall prevalence of glaucoma was 58.5% and was similar for both genes (53.3% for FOXC1 vs 60.9% for PITX2, P=0.59), however, the median age at glaucoma diagnosis was significantly lower in FOXC1 (6.0±13.0 years) compared with PITX2 carriers (18.0±10.6 years, P=0.04). The penetrance at 10 years old was significantly lower in PITX2 than FOXC1 carriers (13.0% vs 42.9%, P=0.03) but became comparable at 25 years old (71.4% vs 57.7%, P=0.38). These findings have important implications for the genetic counselling of families affected by Axenfeld-Rieger syndrome, and also suggest that FOXC1 and PITX2 contribute to the genetic architecture of primary glaucoma subtypes.Emmanuelle Souzeau, Owen M Siggs, Tiger Zhou, Anna Galanopoulos, Trevor Hodson, Deepa Taranath, Richard A Mills, John Landers, John Pater, James E Smith, James E Elder, Julian L Rait, Paul Giles, Vivek Phakey, Sandra E Staffieri, Lisa S Kearns, Andrew Dubowsky, David A Mackey, Alex W Hewitt, Jonathan B Ruddle, Kathryn P Burdon and Jamie E Crai

Uterine Dysfunction in Biglycan and Decorin Deficient Mice Leads to Dystocia during Parturition

Cesarean birth rates are rising. Uterine dysfunction, the exact mechanism of which is unknown, is a common indication for Cesarean delivery. Biglycan and decorin are two small leucine-rich proteoglycans expressed in the extracellular matrix of reproductive tissues and muscle. Mice deficient in biglycan display a mild muscular dystrophy, and, along with mice deficient in decorin, are models of Ehlers-Danlos Syndrome, a connective tissue anomaly associated with uterine rupture. As a variant of Ehlers-Danlos Syndrome is caused by a genetic mutation resulting in abnormal biglycan and decorin secretion, we hypothesized that biglycan and decorin play a role in uterine function. Thus, we assessed wild-type, biglycan, decorin and double knockout pregnancies for timing of birth and uterine function. Uteri were harvested at embryonic days 12, 15 and 18. Nonpregnant uterine samples of the same genotypes were assessed for tissue failure rate and spontaneous and oxytocin-induced contractility. We discovered that biglycan/decorin mixed double-knockout dams displayed dystocia, were at increased risk of delayed labor onset, and showed increased tissue failure in a predominantly decorin-dependent manner. In vitro spontaneous uterine contractile amplitude and oxytocin-induced contractile force were decreased in all biglycan and decorin knockout genotypes compared to wild-type. Notably, we found no significant compensation between biglycan and decorin using quantitative real time PCR or immunohistochemistry. We conclude that the biglycan/decorin mixed double knockout mouse is a model of dystocia and delayed labor onset. Moreover, decorin is necessary for uterine function in a dose-dependent manner, while biglycan exhibits partial compensatory mechanisms in vivo. Thus, this model is poised for use as a model for testing novel targets for preventive or therapeutic manipulation of uterine dysfunction

Impact of the TCR Signal on Regulatory T Cell Homeostasis, Function, and Trafficking

Signaling through the T cell antigen receptor (TCR) is important for the homeostasis of naïve and memory CD4+ T cells. The significance of TCR signaling in regulatory T (Treg) cells has not been systematically addressed. Using an Ox40-cre allele that is prominently expressed in Treg cells, and a conditional null allele of the gene encoding p56Lck, we have examined the importance of TCR signaling in Treg cells. Inactivation of p56Lck resulted in abnormal Treg homeostasis characterized by impaired turnover, preferential redistribution to the lymph nodes, loss of suppressive function, and striking changes in gene expression. Abnormal Treg cell homeostasis and function did not reflect the involvement of p56Lck in CD4 function because these effects were not observed when CD4 expression was inactivated by Ox40-cre.The results make clear multiple aspects of Treg cell homeostasis and phenotype that are dependent on a sustained capacity to signal through the TCR

Analysis combining correlated glaucoma traits identifies five new risk loci for open-angle glaucoma

Open-angle glaucoma (OAG) is a major cause of blindness worldwide. To identify new risk loci for OAG, we performed a genome-wide association study in 3,071 OAG cases and 6,750 unscreened controls, and meta-analysed the results with GWAS data for intraocular pressure (IOP) and optic disc parameters (the overall meta-analysis sample size varying between 32,000 to 48,000 participants), which are glaucoma-related traits. We identified and independently validated four novel genome-wide significant associations within or near MYOF and CYP26A1, LINC02052 and CRYGS, LMX1B, and LMO7 using single variant tests, one additional locus (C9) using gene-based tests, and two genetic pathways - "response to fluid shear stress" and "abnormal retina morphology" - in pathway-based tests. Interestingly, some of the new risk loci contribute to risk of other genetically-correlated eye diseases including myopia and age-related macular degeneration. To our knowledge, this study is the first integrative study to combine genetic data from OAG and its correlated traits to identify new risk variants and genetic pathways, highlighting the future potential of combining genetic data from genetically-correlated eye traits for the purpose of gene discovery and mappin

Dissecting mammalian immunity through mutation.

Although mutation and natural selection have given rise to our immune system, a well-placed mutation can also cripple it, and within an expanding population we are recognizing more and more cases of single-gene mutations that compromise immunity. These mutations are an ideal tool for understanding human immunology, and there are more ways than ever to measure their physiological effects. There are also more ways to create mutations in the laboratory, and to use these resources to systematically define the function of every gene in our genome. This review focuses on the discovery and creation of mutations in the context of mammalian immunity, with an emphasis on the use of genome-wide chemical and CRISPR/Cas9 mutagenesis to reveal gene function