Functional analysis of ESRP1/2 gene variants and CTNND1 isoforms in orofacial cleft pathogenesis.

Orofacial cleft (OFC) is a common human congenital anomaly. Epithelial-specific RNA splicing regulators ESRP1 and ESRP2 regulate craniofacial morphogenesis and their disruption result in OFC in zebrafish, mouse and humans. Using esrp1/2 mutant zebrafish and murine Py2T cell line models, we functionally tested the pathogenicity of human ESRP1/2 gene variants. We found that many variants predicted by in silico methods to be pathogenic were functionally benign. Esrp1 also regulates the alternative splicing of Ctnnd1 and these genes are co-expressed in the embryonic and oral epithelium. In fact, over-expression of ctnnd1 is sufficient to rescue morphogenesis of epithelial-derived structures in esrp1/2 zebrafish mutants. Additionally, we identified 13 CTNND1 variants from genome sequencing of OFC cohorts, confirming CTNND1 as a key gene in human OFC. This work highlights the importance of functional assessment of human gene variants and demonstrates the critical requirement of Esrp-Ctnnd1 acting in the embryonic epithelium to regulate palatogenesis.

ESRP2-microRNA-122 axis directs the postnatal onset of liver polyploidization and maturation.

Hepatocyte polyploidy and maturity are critical to acquiring specialized liver functions. Multiple intra- and extracellular factors influence ploidy, but how they cooperate temporally to steer liver polyploidization and maturation or how post-transcriptional mechanisms integrate into these paradigms is unknown. Here, we identified an important regulatory hierarchy in which postnatal activation of Epithelial-Splicing-Regulatory-Protein-2 (ESRP2) stimulates biogenesis of liver-specific microRNA (miR-122), thereby facilitating polyploidization, maturation, and functional competence of hepatocytes. By determining transcriptome-wide protein-RNA interactions in vivo and integrating them with single-cell and bulk hepatocyte RNA-seq datasets, we delineate an ESRP2-driven RNA processing program that drives sequential replacement of fetal-to-adult transcript isoforms. Specifically, ESRP2 binds the primary miR-122 host gene transcript to promote its processing/biogenesis. Combining constitutive and inducible ESRP2 gain- and loss-of-function mice models with miR-122 rescue experiments, we demonstrate that timed activation of ESRP2 augments miR-122-driven program of cytokinesis failure, ensuring proper onset and extent of hepatocyte polyploidization.

Anti-Nogo-A Antibody Therapy Improves Functional Outcome Following Traumatic Brain Injury.

BACKGROUND: Traumatic brain injury (TBI) can cause sensorimotor deficits, and recovery is slow and incomplete. There are no effective pharmacological treatments for recovery from TBI, but research indicates potential for anti-Nogo-A antibody (Ab) therapy. This Ab neutralizes Nogo-A, an endogenous transmembrane protein that inhibits neuronal plasticity and regeneration. OBJECTIVE: We hypothesized that anti-Nogo-A Ab treatment following TBI results in disinhibited axonal growth from the contralesional cortex, the establishment of new compensatory neuronal connections, and improved function. METHODS: We modeled TBI in rats using the controlled cortical impact method, resulting in focal brain damage and motor deficits like those observed in humans with a moderate cortical TBI. Rats were trained on the skilled forelimb reaching task and the horizontal ladder rung walking task. They were then given a TBI, targeting the caudal forelimb motor cortex, and randomly divided into 3 groups: TBI-only, TBI + Anti-Nogo-A Ab, and TBI + Control Ab. Testing resumed 3 days after TBI and continued for 8 weeks, when rats received an injection of the anterograde neuronal tracer, biotinylated dextran amine (BDA), into the corresponding area contralateral to the TBI. RESULTS: We observed significant improvement in rats that received anti-Nogo-A Ab treatment post-TBI compared to controls. Analysis of BDA-positive axons revealed that anti-Nogo-A Ab treatment resulted in cortico-rubral plasticity to the deafferented red nucleus. Conclusions. Anti-Nogo-A Ab treatment may improve functional recovery via neuronal plasticity to brain areas important for skilled movements, and this treatment shows promise to improve outcomes in humans who have suffered a TBI.

Case report and review of literature: Isolated intramedullary spinal neurocysticercosis.

Background: Cases of isolated intramedullary spinal neurocysticercosis are extremely rare. Only 25 cases have been reported before 2022. Due to its rarity, the diagnosis of spinal neurocysticercosis may be missed. Case presentation: We describe a 37-year-old female patient who developed back pain and lower extremity weakness and was found to have an intramedullary thoracic spine cystic lesion. She was taken to the operating room for resection of the lesion. Pathology revealed a larval cyst wall consistent with neurocysticercosis. The patient was started on albendazole and dexamethasone. Her exam improved post-operatively, and she was able to ambulate with minimal difficulty at the time of follow up. Conclusion: The case provides insights on the diagnosis and treatment of isolated intramedullary spinal neurocysticercosis. Review of the literature suggests that combined surgical and medical intervention results in significant improvement in the patient's neurological exam, and decreases morbidity associated with the disease. We propose a treatment paradigm for this rare manifestation of neurocysticercosis.

The global Protein-RNA interaction map of ESRP1 defines a post-transcriptional program that is essential for epithelial cell function.

The epithelial splicing regulatory proteins, ESRP1 and ESRP2, are essential for mammalian development through the regulation of a global program of alternative splicing of genes involved in the maintenance of epithelial cell function. To further inform our understanding of the molecular functions of ESRP1, we performed enhanced crosslinking immunoprecipitation coupled with high-throughput sequencing (eCLIP) in epithelial cells of mouse epidermis. The genome-wide binding sites of ESRP1 were integrated with RNA-Seq analysis of alterations in splicing and total gene expression that result from epidermal ablation of Esrp1 and Esrp2. These studies demonstrated that ESRP1 functions in splicing regulation occur primarily through direct binding in a position-dependent manner to promote either exon inclusion or skipping. In addition, we also identified widespread binding of ESRP1 in 3' and 5' untranslated regions (UTRs) of genes involved in epithelial cell function, suggesting that its post-transcriptional functions extend beyond splicing regulation.

The germ cell-specific RNA binding protein RBM46 is essential for spermatogonial differentiation in mice.

Control over gene expression is exerted, in multiple stages of spermatogenesis, at the post-transcriptional level by RNA binding proteins (RBPs). We identify here an essential role in mammalian spermatogenesis and male fertility for 'RNA binding protein 46' (RBM46). A highly evolutionarily conserved gene, Rbm46 is also essential for fertility in both flies and fish. We found Rbm46 expression was restricted to the mouse germline, detectable in males in the cytoplasm of premeiotic spermatogonia and meiotic spermatocytes. To define its requirement for spermatogenesis, we generated Rbm46 knockout (KO, Rbm46-/-) mice; although male Rbm46-/- mice were viable and appeared grossly normal, they were infertile. Testes from adult Rbm46-/- mice were small, with seminiferous tubules containing only Sertoli cells and few undifferentiated spermatogonia. Using genome-wide unbiased high throughput assays RNA-seq and 'enhanced crosslinking immunoprecipitation' coupled with RNA-seq (eCLIP-seq), we discovered RBM46 could bind, via a U-rich conserved consensus sequence, to a cohort of mRNAs encoding proteins required for completion of differentiation and subsequent meiotic initiation. In summary, our studies support an essential role for RBM46 in regulating target mRNAs during spermatogonia differentiation prior to the commitment to meiosis in mice.

Evaluation of Sagittal Spinopelvic Balance in Spinal Cord Stimulator Patients.

OBJECTIVE: Spinal cord stimulation (SCS) has become a popular nonopioid pain intervention. However, the treatment failure rate for SCS remains significantly high and many of these patients have poor sagittal spinopelvic balance, which has been found to correlate with increased pain and decreased quality of life. The purpose of this study was to determine if poor sagittal alignment is correlated with SCS treatment failure. MATERIALS AND METHODS: Comparative retrospective analysis was performed between two cohorts of patients who had undergone SCS placement, those who had either subsequent removal of their SCS system (representing a treatment failure cohort) and those that underwent generator replacement (representing a successful treatment cohort). The electronic medical record was used to collect demographic and surgical characteristics, which included radiographic measurements of lumbar lordosis (LL), pelvic incidence (PI), pelvic tilt (PT), and sacral slope (SS). Also included were data on pain medication usage including opioid and nonopioid therapies. RESULTS: Eighty-one patients met inclusion criteria, 31 had complete removal, and 50 had generator replacements. Measurement of sagittal balance parameters demonstrated that many patients had poor alignment, with 34 outside normal range for LL (10 vs 24 in removal and replacement cohorts, respectively), 30 for PI (12 [38.7%] vs 18 [36.0%]), 46 for PT (18 [58.1%] vs 28 [56.0%]), 38 for SS (18 [58.1%] vs 20 [40.0%]), and 39 for PI-LL mismatch (14 [45.2%] vs 25 [50.0%]). There were no significant differences in sagittal alignment parameters between the two cohorts. CONCLUSIONS: This retrospective cohort analysis of SCS patients did not demonstrate any relationship between poor sagittal alignment and failure of SCS therapy. Further studies of larger databases should be performed to determine how many patients ultimately go on to have additional structural spinal surgery after failure of SCS and whether or not those patients go on to have positive outcomes.

Intraoperative Assessment of Coronal Balance with Long Cassette Radiographs in Adult Thoracolumbar Deformity Correction.

BACKGROUND: To describe the use of intraoperative anteroposterior long cassette radiographs (APLCRs) to guide kickstand rod application in adult spinal deformity. METHODS: A retrospective chart review was performed to identify patients with adult thoracolumbar and coronal plane deformity undergoing open segmental decompression with spinopelvic fixation and deformity correction between October 2017 and June 2019 who had APLCRs after rod placement. In patients with persistent intraoperative coronal deviations, a kickstand rod was placed. This supplemental rod was anchored to an iliac screw and to the construct via a pair of side-to-side connectors. A distractor was expended between a vice grip plyer on the kickstand and side-to-side connector to apply a lateralizing force to reduce the degree of deviation. RESULTS: Of 15 patients who underwent T3-ilium fusion with spinal deformity correction with intraoperative APLCRs, 7 underwent kickstand placement. Mean preoperative coronal deviation was similar between cohorts (4.3 cm vs. 2.2 cm, P = 0.09), but was greater intraoperatively in the kickstand cohort (4.3 cm vs. 0.6 cm, P < 0.001). Postoperative coronal deviation was similar between groups (2.1 cm vs. 1.8 cm, P = 0.37). Preoperative fractional lumbar curve was significantly greater in patients requiring a kickstand (23° vs. 35°, P = 0.02), but the major thoracolumbar curve was similar between groups (43° vs. 35°, P = 0.14). CONCLUSIONS: Intraoperative APLCRs can help guide application of a kickstand rod in adult thoracolumbar deformity correction. Patients with a greater fractional lumbar curve may derive greater benefit of kickstand usage, independent of major curve magnitude.

"The Circle" Predicts Extent of Fusion for Surgical Correction of Cervical Spinal Kyphotic Deformities: Proof of Concept.

BACKGROUND: Correction of kyphotic deformities of the spine is a common problem faced by spine surgeons. Unfortunately, there are no clear published guidelines available regarding the extent of spinal fusion required to achieve and maintain lasting deformity correction. We aim to share a set of novel radiographic parameters ("the Circle") that can be used as a guideline for determining the extent of fusion required in surgical correction of spinal kyphotic deformity. METHODS: A Google forms survey was distributed among spine surgeons and trainees to evaluate differences in recommended extent of posterior-approach fusions for cervical spinal kyphotic deformities before and after introduction to the Circle. Extent of fusion before and after use of the Circle were qualitatively and quantitatively analyzed. Data were anonymized and stored in a secure database. RESULTS: Twenty-seven neurosurgical attendings (n = 14), residents (n = 9), and fellows (n = 3) responded to the survey. Variance between predicted upper and lower instrumented vertebrae, and length of construct, was statistically significantly decreased after application of the Circle in almost all cases. Respondents rated the ease of use of the Circle an average of 4.2 out of 5 (5 = the most ease). The majority of participants (92 [6%]; n = 25 of 27) stated that they would or would likely use the Circle as a radiographic tool in the surgical planning for correction of cervical spinal kyphotic deformities in the future. CONCLUSIONS: The Circle is a novel set of radiographic parameters that may be used to educate and guide surgical plans and extent of fusion when aiming to correct spinal kyphotic deformities.

Surgical correction of pediatric spinal deformities with coexisting intraspinal pathology: A case report and literature review.

BACKGROUND: Surgical correction of spinal deformities with coexisting intraspinal pathology (SDCIP) requires special consideration to minimize risks of further injury to an already abnormal spinal cord. However, there is a paucity of literature on this topic. Here, the authors present a pediatric patient with a residual pilocytic astrocytoma and syringomyelia who underwent surgical correction of progressive postlaminectomy kyphoscoliosis. Techniques employed are compared to those in the literature to compile a set of guidelines for surgical correction of SDCIP. METHODS: A systematic MEDLINE search was conducted using the following keywords; "pediatric," "spinal tumor resection," "deformity correction," "postlaminectomy," "scoliosis correction," "intraspinal pathology," "tethered cord," "syringomyelia," or "diastematomyelia." Recommendations for surgical technique for pediatric SDCIP correction were reviewed. RESULTS: The presented case demonstrates recommendations that primarily compressive forces on the convexity of the coronal curve should be used when performing in situ correction of SDCIP. Undercorrection is favored to minimize risks of traction on the abnormal spinal cord. The literature yielded 13 articles describing various intraoperative techniques. Notably, seven articles described use of compressive forces on the convex side of the deformity as the primary mode of correction, while only five articles provided recommendations on how to safely and effectively surgically correct SDCIP. CONCLUSION: The authors demonstrated with their case analysis and literature review that there are no clear current guidelines regarding the safe and effective techniques for in situ correction and fusion for the management of pediatric SDCIP.

Improved Functional Outcome After Peripheral Nerve Stimulation of the Impaired Forelimb Post-stroke.

Lack of blood flow to the brain, i.e., ischemic stroke, results in loss of nerve cells and therefore loss of function in the effected brain regions. There is no effective treatment to improve lost function except restoring blood flow within the first several hours. Rehabilitation strategies are widely used with limited success. The purpose of this study was to examine the effect of electrical stimulation on the impaired upper extremity to improve functional recovery after stroke. We developed a rodent model using an electrode cuff implant onto a single peripheral nerve (median nerve) of the paretic forelimb and applied daily electrical stimulation. The skilled forelimb reaching test was used to evaluate functional outcome after stroke and electrical stimulation. Anterograde axonal tracing from layer V pyramidal neurons with biotinylated dextran amine was done to evaluate the formation of new neuronal connections from the contralesional cortex to the deafferented spinal cord. Rats receiving electrical stimulation on the median nerve showed significant improvement in the skilled forelimb reaching test in comparison with stroke only and stroke with sham stimulation. Rats that received electrical stimulation also exhibited significant improvement in the latency to initiate adhesive removal from the impaired forelimb, indicating better sensory recovery. Furthermore, axonal tracing analysis showed a significant higher midline fiber crossing index in the cervical spinal cord of rats receiving electrical stimulation. Our results indicate that direct peripheral nerve stimulation leads to improved sensorimotor recovery in the stroke-impaired forelimb, and may be a useful approach to improve post-stroke deficits in human patients.

Epigenome-wide association study of kidney function identifies trans-ethnic and ethnic-specific loci.

BACKGROUND: DNA methylation (DNAm) is associated with gene regulation and estimated glomerular filtration rate (eGFR), a measure of kidney function. Decreased eGFR is more common among US Hispanics and African Americans. The causes for this are poorly understood. We aimed to identify trans-ethnic and ethnic-specific differentially methylated positions (DMPs) associated with eGFR using an agnostic, genome-wide approach. METHODS: The study included up to 5428 participants from multi-ethnic studies for discovery and 8109 participants for replication. We tested the associations between whole blood DNAm and eGFR using beta values from Illumina 450K or EPIC arrays. Ethnicity-stratified analyses were performed using linear mixed models adjusting for age, sex, smoking, and study-specific and technical variables. Summary results were meta-analyzed within and across ethnicities. Findings were assessed using integrative epigenomics methods and pathway analyses. RESULTS: We identified 93 DMPs associated with eGFR at an FDR of 0.05 and replicated 13 and 1 DMPs across independent samples in trans-ethnic and African American meta-analyses, respectively. The study also validated 6 previously published DMPs. Identified DMPs showed significant overlap enrichment with DNase I hypersensitive sites in kidney tissue, sites associated with the expression of proximal genes, and transcription factor motifs and pathways associated with kidney tissue and kidney development. CONCLUSIONS: We uncovered trans-ethnic and ethnic-specific DMPs associated with eGFR, including DMPs enriched in regulatory elements in kidney tissue and pathways related to kidney development. These findings shed light on epigenetic mechanisms associated with kidney function, bridging the gap between population-specific eGFR-associated DNAm and tissue-specific regulatory context.

An Irf6-Esrp1/2 regulatory axis controls midface morphogenesis in vertebrates.

Irf6 and Esrp1 are important for palate development across vertebrates. In zebrafish, we found that irf6 regulates the expression of esrp1 We detailed overlapping Irf6 and Esrp1/2 expression in mouse orofacial epithelium. In zebrafish, irf6 and esrp1/2 share expression in periderm, frontonasal ectoderm and oral epithelium. Genetic disruption of irf6 and esrp1/2 in zebrafish resulted in cleft of the anterior neurocranium. The esrp1/2 mutant also developed cleft of the mouth opening. Lineage tracing of cranial neural crest cells revealed that the cleft resulted not from migration defect, but from impaired chondrogenesis. Analysis of aberrant cells within the cleft revealed expression of sox10, col1a1 and irf6, and these cells were adjacent to krt4+ and krt5+ cells. Breeding of mouse Irf6; Esrp1; Esrp2 compound mutants suggested genetic interaction, as the triple homozygote and the Irf6; Esrp1 double homozygote were not observed. Further, Irf6 heterozygosity reduced Esrp1/2 cleft severity. These studies highlight the complementary analysis of Irf6 and Esrp1/2 in mouse and zebrafish, and identify a unique aberrant cell population in zebrafish expressing sox10, col1a1 and irf6 Future work characterizing this cell population will yield additional insight into cleft pathogenesis.

Surgical management of atlantoaxial dislocation and cervical spinal cord injury in craniopagus twins.

A case of cervical spinal cord injury in 12-year-old angular craniopagus twins is presented, with a description of the planning and execution of surgical treatment along with subsequent clinical outcome. The injury occurred following a fall from a standing position, resulting in quadriparesis in one of the twins. Imaging revealed severe craniocervical stenosis resulting from a C1-2 dislocation, and T2-weighted hyperintensity of the cervical spinal cord. After custom halo fixation was obtained, a posterior approach was utilized to decompress and instrument the occiput, cervical, and upper thoracic spine with intraoperative reduction of the dislocation. Early neurological improvement was noted during the acute postoperative phase, and 27 months of follow-up demonstrated intact instrumentation with continued neurological improvement to near baseline. The complexity of managing such an injury, inclusive of the surgical, anesthetic, biomechanical, and ethical considerations, is described in detail.

Clinical Phenotypes of Atopy and Asthma in COPD: A Meta-analysis of SPIROMICS and COPDGene.

BACKGROUND: Little is known about the concordance of atopy with asthma COPD overlap. Among individuals with COPD, a better understanding of the phenotypes characterized by asthma overlap and atopy is needed to better target therapies. RESEARCH QUESTION: What is the overlap between atopy and asthma status among individuals with COPD, and how are categories defined by the presence of atopy and asthma status associated with clinical and radiologic phenotypes and outcomes in the Genetic Epidemiology of COPD Study (COPDGene) and Subpopulation and Intermediate Outcome Measures in COPD Study (SPIROMICS)? STUDY DESIGN AND METHODS: Four hundred three individuals with COPD from SPIROMICS and 696 individuals from COPDGene with data about specific IgEs to 10 common allergens and mixes (simultaneous assessment of combination of allergens in similar category) were included. Comparison groups were defined by atopic and asthma status (neither, atopy alone, atopic asthma, nonatopic asthma, with atopy defined as any positive specific IgE (≥0.35 KU/L) to any of the 10 allergens or mixes and asthma defined as self-report of doctor-diagnosed current asthma). Multivariable regression analyses (linear, logistic, and zero inflated negative binomial where appropriate) adjusted for age, sex, race, lung function, smoking status, pack-years smoked, and use of inhaled corticosteroids were used to determine characteristics of groups and relationship with outcomes (exacerbations, clinical outcomes, CT metrics) separately in COPDGene and SPIROMICS, and then adjusted results were combined using meta-analysis. RESULTS: The prevalence of atopy was 35% and 36% in COPD subjects from SPIROMICS and COPDGene, respectively, and less than 50% overlap was seen between atopic status with asthma in both cohorts. In meta-analysis, individuals with nonatopic asthma had the most impaired symptom scores (effect size for St. George's Respiratory Questionnaire total score, 4.2; 95% CI, 0.4-7.9; effect size for COPD Assessment Test score, 2.8; 95% CI, 0.089-5.4), highest risk for exacerbations (incidence rate ratio, 1.41; 95% CI, 1.05-1.88) compared with the group without atopy or asthma. Those with atopy and atopic asthma were not at increased risk for adverse outcomes. INTERPRETATION: Asthma and atopy had incomplete overlap among former and current smokers with COPD in COPDGene and SPIROMICS. Nonatopic asthma was associated with adverse outcomes and exacerbation risk in COPD, whereas groups having atopy alone and atopic asthma had less risk.

Cleft lip and cleft palate in Esrp1 knockout mice is associated with alterations in epithelial-mesenchymal crosstalk.

Cleft lip is one of the most common human birth defects. However, there remain a limited number of mouse models of cleft lip that can be leveraged to characterize the genes and mechanisms that cause this disorder. Crosstalk between epithelial and mesenchymal cells underlies formation of the face and palate, but the basic molecular events mediating this crosstalk remain poorly understood. We previously demonstrated that mice lacking the epithelial-specific splicing factor Esrp1 have fully penetrant bilateral cleft lip and palate. In this study, we further investigated the mechanisms leading to cleft lip as well as cleft palate in both existing and new Esrp1 mutant mouse models. These studies included a detailed transcriptomic analysis of changes in ectoderm and mesenchyme in Esrp1-/- embryos during face formation. We identified altered expression of genes previously implicated in cleft lip and/or palate, including components of multiple signaling pathways. These findings provide the foundation for detailed investigations using Esrp1 mutant disease models to examine gene regulatory networks and pathways that are essential for normal face and palate development - the disruption of which leads to orofacial clefting in human patients.

Tarik Syndrome: Reversible Postoperative Blindness Secondary to Occipital Seizures.

BACKGROUND: Postoperative blindness is a devastating surgical complication. Although usually associated with prolonged cardiac and prone spinal operations, it may follow other procedures as well. Postoperative blindness is most commonly caused by a vascular etiology, but it can more rarely be caused by status epilepticus. We have previously reported a case of this phenomenon following a staged spinal deformity surgery. CASE DESCRIPTION: Here we report 2 additional cases following a skull base procedure and a single stage lumbar spine surgery. In all instances, rapid recognition that the patients' blindness was due to occipital seizures resulted in acute antiepileptiform treatment and full restoration of vision. CONCLUSIONS: Although a rare phenomenon, this syndrome, first recognized and described by Tarik F. Ibrahim, should be considered in any patient with postoperative visual impairment.

Transverse myelitis associated with yellow fever vaccination.

Yellow fever is a mosquito-borne viral hemorrhagic fever endemic to sub-Saharan Africa and South America associated with significant morbidity and mortality. The use of a live attenuated vaccine can prevent yellow fever, but vaccine-associated neurologic disease has been reported and is a safety concern. We present the case of a previously healthy 35-year-old active-duty man who received the yellow fever vaccine prior to deployment and subsequently developed progressive neurologic dysfunction consistent with transverse myelitis.

Cerebellar Hemorrhage Following an Uncomplicated Lumbar Spine Surgery: Case Report.

Remote cerebellar hemorrhage is rare but potentially fatal complication of cranial and spinal surgeries. The pathophysiology of this condition following spinal surgery is thought to be related to venous bleeding from cerebellar sagging and cerebrospinal fluid (CSF) hypotension. Most reported cases in the literature following spinal surgery involve intraoperative CSF leakage. We present a case of remote cerebellar hemorrhage following uncomplicated lumbar spinal decompression and fusion without CSF leakage.