Neuropathy target esterase activity defines phenotypes among PNPLA6 disorders.

Biallelic pathogenic variants in the PNPLA6 gene cause a broad spectrum of disorders leading to gait disturbance, visual impairment, anterior hypopituitarism and hair anomalies. PNPLA6 encodes neuropathy target esterase (NTE), yet the role of NTE dysfunction on affected tissues in the large spectrum of associated disease remains unclear. We present a systematic evidence-based review of a novel cohort of 23 new patients along with 95 reported individuals with PNPLA6 variants that implicate missense variants as a driver of disease pathogenesis. Measuring esterase activity of 46 disease-associated and 20 common variants observed across PNPLA6-associated clinical diagnoses unambiguously reclassified 36 variants as pathogenic and 10 variants as likely pathogenic, establishing a robust functional assay for classifying PNPLA6 variants of unknown significance. Estimating the overall NTE activity of affected individuals revealed a striking inverse relationship between NTE activity and the presence of retinopathy and endocrinopathy. This phenomenon was recaptured in vivo in an allelic mouse series, where a similar NTE threshold for retinopathy exists. Thus, PNPLA6 disorders, previously considered allelic, are a continuous spectrum of pleiotropic phenotypes defined by an NTE genotype:activity:phenotype relationship. This relationship, and the generation of a preclinical animal model, pave the way for therapeutic trials, using NTE as a biomarker.

Optic neuropathy from hypovitaminosis A in a series of children with severe dietary restrictions.

AIM: Hypovitaminosis A is a leading cause of preventable childhood blindness, especially in developing nations. Vitamin A is a fat-soluble essential micronutrient that serves vital functions in the visual system and in regulating bone resorption. We report on a series of four children with mixed nutritional and compressive optic neuropathy and provide a review of the literature. METHODS: A retrospective observational study of four males (ages 9-12), three with autism spectrum disorder who presented with loss of vision and multiple vitamin deficiencies including hypovitaminosis A. RESULTS: Patients presented with unexplained visual loss or a change in visual behaviour. All patients had severely restricted diet comprising of predominantly carbohydrates. Two of the four cases demonstrated optic nerve pallor at initial presentation with marked optic atrophy developing in all patients over time. Electrophysiology available in two patients demonstrated optic nerve dysfunction with preserved retinal function. Extensive investigations revealed profound deficiency in multiple vitamins including vitamin A (<0.1-0.2 µmol/L, normal = 0.9-1.7 µmol/L). Three patients also had low vitamin B12 (90-111 pmol/L, normal = 170-800 pmol/L) with normal folate. All four cases had radiological evidence of skull base thickening indicative of low vitamin A. Genetic testing did not find any relevant pathogenic variants. CONCLUSIONS: Hypovitaminosis A is a crucial form of nutritional deprivation that results in significant visual loss with potential hyperostosis and optic nerve compression exacerbating nutritional optic neuropathy. Additional micronutrient deficiencies usually co-exist and may contribute. Extra vigilance in vitamin replacement is required of clinicians with patients with autism who have restricted diets.

Optic Atrophy From Retrograde Transsynaptic Axonal Degeneration Following Pediatric Brain Injury.

BACKGROUND: The patterns of optic atrophy due to retrograde transsynaptic degeneration (RTSD) have not been well characterized in children. This study aimed to characterize optic atrophy in pediatric patients with focal intracerebral lesions. METHODS: A retrospective review of children with optic atrophy and focal intracerebral lesions was conducted. Ophthalmic data were recorded, including visual acuity, color vision, formal automated visual fields and optical coherence tomography (OCT) of the peripapillary retinal nerve fiber layer (pRNFL) and ganglion cell layer. RESULTS: Six patients (83.33% male) were included. The mean visual acuity (VA) of all eyes was 0.30 logMAR (20/40 Snellen), with no significant difference in the mean logMAR VA in the ipsilateral eye to the location of the lesion compared with the contralateral eye (0.30 vs 0.30, P = 1.000). Color vision (available in 5 patients) was normal in 2, mildly reduced in one and markedly reduced in 2. Bitemporal optic disc pallor was observed in 5 out of 6 patients. OCT data revealed that pRNFL thickness was most significantly diminished in the temporal (95% CI: -44.71 to -14.18 µm, P = 0.0021), inferotemporal (95% CI: -75.06 to -5.17 µm, P = 0.0294), and superotemporal (95% CI: -76.82 to -18.51 µm, P = 0.0055) sectors. Average pRNFL thickness was significantly reduced compared with normative data in both the ipsilateral (95% CI: -40.76 to -11.69 µm, P = 0.0003) and the contralateral eye (95% CI: -38.46 to -5.83 µm, P = 0.0063). When only nasal and temporal data were analyzed, mean pRNFL thickness was still diminished compared with normative data (95% CI: -33.01 to -9.77 µm, P = 0.0012). CONCLUSIONS: Children presenting with optic atrophy, particularly with bitemporal optic atrophy, should have neuroimaging to exclude any underlying serious intracranial pathology.

Planting the Seed for Blood Pressure Control: The Role of Plant-Based Nutrition in the Management of Hypertension.

PURPOSE OF REVIEW: Hypertension results in significant morbidity, mortality, and healthcare expenditures. Fortunately, it is largely preventable and treatable by implementing dietary interventions, though these remain underutilized. Here, we aim to explore the role of healthy dietary patterns in hypertension management and describe approaches for busy clinicians to address nutrition effectively and efficiently with patients. RECENT FINDINGS: DASH, Mediterranean, vegetarian, and vegan diets that include minimally processed, plant-based foods as core elements have consistently shown positive effects on hypertension. Recommendations that distill the most healthful components of these diets can significantly impact patient outcomes. Clinicians can harness evidence-based dietary assessment and counseling tools to implement and support behavioral changes, even during brief office visits. Healthful plant-based dietary patterns can often effectively prevent and treat hypertension. Clinicians may help improve patient outcomes by discussing evidence-based nutrition with their patients. Future work to promote infrastructural change that supports incorporating evidence-based nutrition into medical education, clinical care, and society at large can support these efforts.

Chlamydia trachomatis conjunctivitis in the pre-pubertal child.

Much is reported in the literature about the transmission and presentation of Chlamydia trachomatis conjunctival infection in the neonate; however, there is a paucity of information available on infection in the older pre-pubertal child (>3 years of age). We present the case of a 7-year-old girl, referred for assessment at the sexual assault referral centre following the diagnosis of unilateral C. trachomatis conjunctivitis. This child underwent a rigorous multiagency child protection process, with input from medical professionals, social services and the police to investigate the possibility of child sexual abuse (CSA). However, a group consensus was reached that non-sexual close contact transfer of C. trachomatis from the mother was the most likely mode of transmission and cause of infection. We aim to take the reader through the complex path to this conclusion, the approach to sexually transmitted infections and potential CSA and what is currently known about chlamydial conjunctivitis in children beyond the neonatal period.

Periosteal fixation of the medial and lateral recti for large-angle incomitant exotropia.

PURPOSE: To describe the evolution of a surgical technique for the correction of large-angle incomitant exodeviations. METHODS: Retrospective review of an interventional case series from 2005 to 2019 in a single centre, with analysis of surgical procedure, prism diopter (PD) deviations and complications. RESULTS: Thirty-one patients underwent surgery at an average age of 42 years (range 4-75 years) for minimal medial rectus function, mostly from oculomotor nerve palsy (23/31; 74%). The mean pre-operative exodeviation was 75 PD (range 30-200PD). Sixteen patients (52%) had undergone previous strabismus surgery. Thirty-eight operations were performed in which the medial rectus insertion was anchored to the periosteum of the posterior lacrimal crest via a retrocaruncular transconjunctival approach. The ipsilateral lateral rectus (LR) was disinserted and fixed to lateral orbital tissue in 29/38 (76%) operations, injected with botulinum toxin in 5, recessed in 2 and had already undergone maximal LR recession in 2. In all but the first 8 operations, temporary limbal sutures were passed through the eyelids to maximally adduct the globe post-operatively. At last follow-up (mean 24 months; range 2-130), the mean reduction in exodeviation was 49PD (range 10-80) and overall residual deviation was 26PD (range 80PD base-in to 14PD base-out). The 5 LR toxin procedures had a mean reduction of 22PD (range 10-40). Seven patients had persistent diplopia, one a transient corneal erosion and one caruncle suture exposure 4 years after surgery. CONCLUSION: Large-angle exodeviations can be markedly improved by bi-rectus fixation. This approach is both safe and effective and can be performed in complex patients with multiple previous procedures.

Practical and Ethical Considerations in Telehealth: Pitfalls and Opportunities.

Telehealth has been a long-awaited advancement with the potential to improve efficiency, convenience, and quality in healthcare. However, as telehealth becomes integrated into routine clinical care, it is imperative to consider the practical and ethical implications that could undermine or devalue care delivery. The medical profession must ensure that it is implemented judiciously and with robust quality standards, guided by fair and equitable policies that balance patient autonomy with rigorous standards of care and access. Such a system must recognize the opportunity for more patient input as stakeholders to tailor care to their needs and preferences, while also acknowledging the risk of suboptimal care if convenience is prioritized over quality. More studies of optimal care models are needed to integrate data in terms of both stakeholder input and outcomes.

3D Bioprinting using UNIversal Orthogonal Network (UNION) Bioinks.

Three-dimensional (3D) bioprinting is a promising technology to produce tissue-like structures, but a lack of diversity in bioinks is a major limitation. Ideally each cell type would be printed in its own customizable bioink. To fulfill this need for a universally applicable bioink strategy, we developed a versatile, bioorthogonal bioink crosslinking mechanism that is cell compatible and works with a range of polymers. We term this family of materials UNIversal, Orthogonal Network (UNION) bioinks. As demonstration of UNION bioink versatility, gelatin, hyaluronic acid (HA), recombinant elastin-like protein (ELP), and polyethylene glycol (PEG) were each used as backbone polymers to create inks with storage moduli spanning 200 to 10,000 Pa. Because UNION bioinks are crosslinked by a common chemistry, multiple materials can be printed together to form a unified, cohesive structure. This approach is compatible with any support bath that enables diffusion of UNION crosslinkers. Both matrix-adherent human corneal mesenchymal stromal cells and non-matrix-adherent human induced pluripotent stem cell-derived neural progenitor spheroids were printed with UNION bioinks. The cells retained high viability and expressed characteristic phenotypic markers after printing. Thus, UNION bioinks are a versatile strategy to expand the toolkit of customizable materials available for 3D bioprinting.

Mutation in the intracellular chloride channel CLCC1 associated with autosomal recessive retinitis pigmentosa.

We identified a homozygous missense alteration (c.75C>A, p.D25E) in CLCC1, encoding a presumptive intracellular chloride channel highly expressed in the retina, associated with autosomal recessive retinitis pigmentosa (arRP) in eight consanguineous families of Pakistani descent. The p.D25E alteration decreased CLCC1 channel function accompanied by accumulation of mutant protein in granules within the ER lumen, while siRNA knockdown of CLCC1 mRNA induced apoptosis in cultured ARPE-19 cells. TALEN KO in zebrafish was lethal 11 days post fertilization. The depressed electroretinogram (ERG) cone response and cone spectral sensitivity of 5 dpf KO zebrafish and reduced eye size, retinal thickness, and expression of rod and cone opsins could be rescued by injection of wild type CLCC1 mRNA. Clcc1+/- KO mice showed decreased ERGs and photoreceptor number. Together these results strongly suggest that intracellular chloride transport by CLCC1 is a critical process in maintaining retinal integrity, and CLCC1 is crucial for survival and function of retinal cells.

Reflections on New Evidence on Crisis Standards of Care in the COVID-19 Pandemic.

Crisis standards of care have been widely developed by healthcare systems and states in the United States during the COVID-19 pandemic, and in some rare cases have actually been used to allocate medical resources. All publicly available U.S. crisis standards of care with a mechanism for allocating scarce resources make use of the Sequential Organ Failure Assessment (SOFA) score in hopes of assigning scarce resources to those patients who are more likely to survive. We reflect on the growing body of evidence suggesting that the SOFA score has limited accuracy in predicting mortality among patients hospitalized with COVID-19 and that the SOFA score systematically disfavors Black patients. Use of the SOFA score for allocating scarce resources may therefore result in Black patients with equal likelihood of survival being deprived of life-saving medical resources. There is also a risk of injustice for patients with non-COVID-19 diagnoses, for whom the SOFA score may be a more accurate prognostic score, but who might nevertheless be unfairly (de)prioritized when assessed alongside COVID-19 patients using the same scoring system. For these reasons we recommend that the SOFA score not be used for triage purposes during the COVID pandemic, and that a national effort be made to develop and empirically test crisis standards of care in advance of the next public health emergency.

Molecular and phenotypic investigation of a New Zealand cohort of childhood-onset retinal dystrophy.

Inherited retinal diseases are clinically heterogeneous and are associated with nearly 300 different genes. In this retrospective, observational study of a consecutive cohort of 159 patients (134 families) with childhood-onset (<16 years of age) retinal dystrophy, molecular investigations, and in-depth phenotyping were performed to determine key clinical and molecular characteristics. The most common ocular phenotype was rod-cone dystrophy in 40 patients. Leber Congenital Amaurosis, the most severe form of retinal dystrophy, was present in 10 patients, and early onset severe retinal dystrophy in 22 patients. Analysis has so far identified 131 pathogenic or likely pathogenic variants including 22 novel variants. Molecular diagnosis was achieved in 112 of 134 families (83.6%) by NGS gene panel investigation in 60 families, Sanger sequencing in 27 families, and Asper microarray in 25 families. An additional nine variants of uncertain significance were also found including three novel variants. Variants in 36 genes have been identified with the most common being ABCA4 retinopathy in 36 families. Five sporadic retinal dystrophy patients were found to have variants in dominant and X-linked genes (CRX, RHO, RP2, and RPGR) resulting in more accurate genetic counseling of inheritance for these families. Variants in syndromic associated genes including ALMS1, SDCCAG8, and PPT1 were identified in eight families enabling directed systemic care.

Biallelic Mutation of ARHGEF18, Involved in the Determination of Epithelial Apicobasal Polarity, Causes Adult-Onset Retinal Degeneration.

Mutations in more than 250 genes are implicated in inherited retinal dystrophy; the encoded proteins are involved in a broad spectrum of pathways. The presence of unsolved families after highly parallel sequencing strategies suggests that further genes remain to be identified. Whole-exome and -genome sequencing studies employed here in large cohorts of affected individuals revealed biallelic mutations in ARHGEF18 in three such individuals. ARHGEF18 encodes ARHGEF18, a guanine nucleotide exchange factor that activates RHOA, a small GTPase protein that is a key component of tight junctions and adherens junctions. This biological pathway is known to be important for retinal development and function, as mutation of CRB1, encoding another component, causes retinal dystrophy. The retinal structure in individuals with ARHGEF18 mutations resembled that seen in subjects with CRB1 mutations. Five mutations were found on six alleles in the three individuals: c.808A>G (p.Thr270Ala), c.1617+5G>A (p.Asp540Glyfs∗63), c.1996C>T (p.Arg666∗), c.2632G>T (p.Glu878∗), and c.2738_2761del (p.Arg913_Glu920del). Functional tests suggest that each disease genotype might retain some ARHGEF18 activity, such that the phenotype described here is not the consequence of nullizygosity. In particular, the p.Thr270Ala missense variant affects a highly conserved residue in the DBL homology domain, which is required for the interaction and activation of RHOA. Previously, knock-out of Arhgef18 in the medaka fish has been shown to cause larval lethality which is preceded by retinal defects that resemble those seen in zebrafish Crumbs complex knock-outs. The findings described here emphasize the peculiar sensitivity of the retina to perturbations of this pathway, which is highlighted as a target for potential therapeutic strategies.

Comprehensive Rare Variant Analysis via Whole-Genome Sequencing to Determine the Molecular Pathology of Inherited Retinal Disease.

Inherited retinal disease is a common cause of visual impairment and represents a highly heterogeneous group of conditions. Here, we present findings from a cohort of 722 individuals with inherited retinal disease, who have had whole-genome sequencing (n = 605), whole-exome sequencing (n = 72), or both (n = 45) performed, as part of the NIHR-BioResource Rare Diseases research study. We identified pathogenic variants (single-nucleotide variants, indels, or structural variants) for 404/722 (56%) individuals. Whole-genome sequencing gives unprecedented power to detect three categories of pathogenic variants in particular: structural variants, variants in GC-rich regions, which have significantly improved coverage compared to whole-exome sequencing, and variants in non-coding regulatory regions. In addition to previously reported pathogenic regulatory variants, we have identified a previously unreported pathogenic intronic variant in CHM in two males with choroideremia. We have also identified 19 genes not previously known to be associated with inherited retinal disease, which harbor biallelic predicted protein-truncating variants in unsolved cases. Whole-genome sequencing is an increasingly important comprehensive method with which to investigate the genetic causes of inherited retinal disease.

Supplemental 25-Hydroxycholecalciferol Is More Effective than Cholecalciferol in Raising Serum 25-Hydroxyvitamin D Concentrations in Older Adults.

BACKGROUND: There are few studies directly comparing the pharmacokinetics of 25-hydroxycholecalciferol [25(OH)D3] to cholecalciferol (D3). OBJECTIVES: The primary objectives were to compare the effectiveness of D3 and 25(OH)D3 in raising 25-hydroxyvitamin D [25(OH)D] serum concentrations and achieving steady state. METHODS: This was a randomized, double-blind, active comparator trial of 91 participants (53 females, 38 males), aged 63.3 ± 7.9 y. 25(OH)D3 (10, 15, and 20 µg) and D3 (20 µg) were dosed daily for 6 mo followed by 6 mo of washout. Frequent measurements of serum 25(OH)D were performed. Pharmacokinetic parameters were fitted for each individual and the treatment average was modeled with linear regression using the individual baseline level, sex, and gender as covariates. RESULTS: Mean baseline 25(OH)D was similar in all groups (47.1-49.5 nmol/L). Increases in 25(OH)D to steady state were higher in the 25(OH)D3 groups than in the D3 group [least squares (LS) means (95% CI): 50.1 (43.3-58.0), 72.5 (64.3-81.7), 97.4 (86.6-109.6) nmol/L in 10, 15, and 20 µg/d and 38.7 (33.1-45.2) nmol/L in the D3 group; P = 0.0173, P < 0.0001, P < 0.0001]. The rate to reach steady state was similar in all groups, but the time to reach 25(OH)D concentrations of 75 nmol/L was faster in the higher-dosed 25(OH)D3 groups than in the D3 group (7 and 10 d compared with 40 d, P < 0.0001 and P < 0.0001 for 15 and 20 µg/d). The rate of elimination was 59-109% higher in the 25(OH)D3 groups than in the D3 group. The area under the curve (AUC)/µg dose demonstrated that 25(OH)D3 was 3 times as effective as D3 at raising 25(OH)D concentrations. CONCLUSIONS: 25(OH)D3 is ∼3 times as effective as D3 at raising 25(OH)D concentrations. Once supplementation is discontinued, the elimination rate of 25(OH)D3 is faster than D3. This trial was registered at clinicaltrials.gov as NCT02333682.

Triage and justice in an unjust pandemic: ethical allocation of scarce medical resources in the setting of racial and socioeconomic disparities.

Shortages of life-saving medical resources caused by COVID-19 have prompted hospitals, healthcare systems, and governmentsto develop crisis standards of care, including 'triage protocols' to potentially ration medical supplies during the public health emergency. At the same time, the pandemic has highlighted and exacerbated racial, ethnic, and socioeconomic health disparities that together constitute a form of structural racism. These disparities pose a critical ethical challenge in developing fair triage systems that will maximize lives saved without perpetuating systemic inequities. Here we review alternatives to 'utilitarian' triage, including first-come first-served, egalitarian, and prioritarian systems of allocating scarce medical resources. We assess the comparative advantages and disadvantages of these allocation schemes. Ultimately, we argue that while triage protocols should not exacerbate disparities, they are not an adequate mechanism for redressing systemic health inequities. Entrenched health disparities must be addressed through broader social change.

Developing a Triage Protocol for the COVID-19 Pandemic: Allocating Scarce Medical Resources in a Public Health Emergency.

The coronavirus disease-2019 (COVID-19) has caused shortages of life-sustaining medical resources, and future waves of the virus may cause further scarcity. The Yale New Haven Health System developed a triage protocol to allocate scarce medical resources during the COVID-19 pandemic, with the primary goal of saving the most lives possible, and a secondary goal of making triage assessments and decisions consistent, transparent, and fair. We outline the process of developing the triage protocol, summarize the protocol itself, and discuss the major ethical challenges encountered, along with our answers to these challenges. These challenges include (1) the role of age and chronic comorbidities; (2) evaluating children and pregnant patients; (3) racial, ethnic, and socioeconomic disparities in health; (4) prioritization of healthcare workers; and (5) balancing clinical judgment versus protocolized assessments. We conclude with a review of the limitations of our protocol and the lessons learned. We hope that a robust public discussion of such protocols and the ethical challenges that they raise will result in the fairest possible processes, less need for triage, and more lives saved during future waves of the COVID-19 pandemic and similar public health emergencies.

Mutations in REEP6 Cause Autosomal-Recessive Retinitis Pigmentosa.

Retinitis pigmentosa (RP) is the most frequent form of inherited retinal dystrophy. RP is genetically heterogeneous and the genes identified to date encode proteins involved in a wide range of functional pathways, including photoreceptor development, phototransduction, the retinoid cycle, cilia, and outer segment development. Here we report the identification of biallelic mutations in Receptor Expression Enhancer Protein 6 (REEP6) in seven individuals with autosomal-recessive RP from five unrelated families. REEP6 is a member of the REEP/Yop1 family of proteins that influence the structure of the endoplasmic reticulum but is relatively unstudied. The six variants identified include three frameshift variants, two missense variants, and a genomic rearrangement that disrupts exon 1. Human 3D organoid optic cups were used to investigate REEP6 expression and confirmed the expression of a retina-specific isoform REEP6.1, which is specifically affected by one of the frameshift mutations. Expression of the two missense variants (c.383C>T [p.Pro128Leu] and c.404T>C [p.Leu135Pro]) and the REEP6.1 frameshift mutant in cultured cells suggest that these changes destabilize the protein. Furthermore, CRISPR-Cas9-mediated gene editing was used to produce Reep6 knock-in mice with the p.Leu135Pro RP-associated variant identified in one RP-affected individual. The homozygous knock-in mice mimic the clinical phenotypes of RP, including progressive photoreceptor degeneration and dysfunction of the rod photoreceptors. Therefore, our study implicates REEP6 in retinal homeostasis and highlights a pathway previously uncharacterized in retinal dystrophy.

Care Labor in VAD Therapy: Some Feminist Concerns.

Ventricular assist device (VAD) care offers a distinctive lens through which we can explore unjust gender norms. This is a resource-intensive intervention, one in which increasingly sophisticated technology brings with it the need for more long-term care. This care work is demanding, involving device maintenance, medication and appointment management, household work, and emotional support. Most patients eligible for receiving VADs are men, so it is not surprising that it is more often women who are responsible for the care of patients with VADs. Still, there is room to question why so much of this labor is expected of and taken on by female caregivers, when it could be shared with male caregivers and even patients themselves. To the extent that gender difference in the distribution of this labor is avoidable and inequitable, it becomes in part a disparity resulting from unjust social norms. In order to unpack some of this injustice, the authors utilize empiric data and theoretical work in feminist ethics to articulate some of the mechanisms of the gender disparity in VAD care labor and to offer communitarian decision-making and redistribution of care labor as potential routes toward greater justice for women with respect to VAD therapy.

Mutations in TUBGCP4 alter microtubule organization via the γ-tubulin ring complex in autosomal-recessive microcephaly with chorioretinopathy.

We have identified TUBGCP4 variants in individuals with autosomal-recessive microcephaly and chorioretinopathy. Whole-exome sequencing performed on one family with two affected siblings and independently on another family with one affected child revealed compound-heterozygous mutations in TUBGCP4. Subsequent Sanger sequencing was performed on a panel of individuals from 12 French families affected by microcephaly and ophthalmic manifestations, and one other individual was identified with compound-heterozygous mutations in TUBGCP4. One synonymous variant was common to all three families and was shown to induce exon skipping; the other mutations were frameshift mutations and a deletion. TUBGCP4 encodes γ-tubulin complex protein 4, a component belonging to the γ-tubulin ring complex (γ-TuRC) and known to regulate the nucleation and organization of microtubules. Functional analysis of individual fibroblasts disclosed reduced levels of the γ-TuRC, altered nucleation and organization of microtubules, abnormal nuclear shape, and aneuploidy. Moreover, zebrafish treated with morpholinos against tubgcp4 were found to have reduced head volume and eye developmental anomalies with chorioretinal dysplasia. In summary, the identification of TUBGCP4 mutations in individuals with microcephaly and a spectrum of anomalies in eye development, particularly photoreceptor anomalies, provides evidence of an important role for the γ-TuRC in brain and eye development.

Missense mutations in the WD40 domain of AHI1 cause non-syndromic retinitis pigmentosa.

BACKGROUND: Recent findings suggesting that Abelson helper integration site 1 (AHI1) is involved in non-syndromic retinal disease have been debated, as the functional significance of identified missense variants was uncertain. We assessed whether AHI1 variants cause non-syndromic retinitis pigmentosa (RP). METHODS: Exome sequencing was performed in three probands with RP. The effects of the identified missense variants in AHI1 were predicted by three-dimensional structure homology modelling. Ciliary parameters were evaluated in patient's fibroblasts, and recombinant mutant proteins were expressed in ciliated retinal pigmented epithelium cells. RESULTS: In the three patients with RP, three sets of compound heterozygous variants were detected in AHI1 (c.2174G>A; p.Trp725* and c.2258A>T; p.Asp753Val, c.660delC; p.Ser221Glnfs*10 and c.2090C>T; p.Pro697Leu, c.2087A>G; p.His696Arg and c.2429C>T; p.Pro810Leu). All four missense variants were present in the conserved WD40 domain of Jouberin, the ciliary protein encoded by AHI1, with variable predicted implications for the domain structure. No significant changes in the percentage of ciliated cells, nor in cilium length or intraflagellar transport were detected. However, expression of mutant recombinant Jouberin in ciliated cells showed a significantly decreased enrichment at the ciliary base. CONCLUSIONS: This report confirms that mutations in AHI1 can underlie autosomal recessive RP. Moreover, it structurally and functionally validates the effect of the RP-associated AHI1 variants on protein function, thus proposing a new genotype-phenotype correlation for AHI1 mutation associated retinal ciliopathies.