Crispant analysis in zebrafish as a tool for rapid functional screening of disease-causing genes for bone fragility.

Heritable fragile bone disorders (FBDs), ranging from multifactorial to rare monogenic conditions, are characterized by an elevated fracture risk. Validating causative genes and understanding their mechanisms remain challenging. We assessed a semi-high throughput zebrafish screening platform for rapid in vivo functional testing of candidate FBD genes.

Bmpr1aa modulates the severity of the skeletal phenotype in an fkbp10-deficient Bruck syndrome zebrafish model.

Rare monogenic disorders often exhibit significant phenotypic variability among individuals sharing identical genetic mutations. Bruck syndrome (BS), a prime example, is characterized by bone fragility and congenital contractures, although with a pronounced variability among family members. BS arises from recessive biallelic mutations in FKBP10 or PLOD2.

Syntaxin 18 Defects in Human and Zebrafish Unravel Key Roles in Early Cartilage and Bone Development.

SNARE proteins comprise a conserved protein family responsible for catalyzing membrane fusion during vesicle traffic. Syntaxin18 (STX18) is a poorly characterized endoplasmic reticulum (ER)-resident t-SNARE. Recently, together with TANGO1 and SLY1, its involvement was shown in ER to Golgi transport of collagen II during chondrogenesis.

Poly (A)-specific ribonuclease deficiency impacts oogenesis in zebrafish.

Poly (A)-specific ribonuclease (PARN) is the most important 3'-5'exonuclease involved in the process of deadenylation, the removal of poly (A) tails of mRNAs. Although PARN is primarily known for its role in mRNA stability, recent studies suggest several other functions of PARN including a role in telomere biology, non-coding RNA maturation, trimming of miRNAs, ribosome biogenesis and TP53 function. Moreover, PARN expression is de-regulated in many cancers, including solid tumours and hematopoietic malignancies.

A tapt1 knock-out zebrafish line with aberrant lens development and impaired vision models human early-onset cataract.

Bi-allelic mutations in the gene coding for human trans-membrane anterior-posterior transformation protein 1 (TAPT1) result in a broad phenotypic spectrum, ranging from syndromic disease with severe skeletal and congenital abnormalities to isolated early-onset cataract. We present here the first patient with a frameshift mutation in the TAPT1 gene, resulting in both bilateral early-onset cataract and skeletal abnormalities, in addition to several dysmorphic features, in this way further expanding the phenotypic spectrum associated with TAPT1 mutations. A tapt1a/tapt1b double knock-out (KO) zebrafish model generated by CRISPR/Cas9 gene editing revealed an early larval phenotype with eye malformations, loss of vision, increased photokinetics and hyperpigmentation, without visible skeletal involvement.

Online Sports Medicine Fellowship Education: The Genesis of a National Program and Year-1 Analysis.

The COVID-19 pandemic has created numerous challenges in all walks of life. One such challenge was the strain and subsequent effects on medical education, including the elimination of in-person learning opportunities. Consequently, in March of 2020, a nationwide Sports Medicine fellowship online education series was developed.

Lrp5 Mutant and Crispant Zebrafish Faithfully Model Human Osteoporosis, Establishing the Zebrafish as a Platform for CRISPR-Based Functional Screening of Osteoporosis Candidate Genes.

Genomewide association studies (GWAS) have improved our understanding of the genetic architecture of common complex diseases such as osteoporosis. Nevertheless, to attribute functional skeletal contributions of candidate genes to osteoporosis-related traits, there is a need for efficient and cost-effective in vivo functional testing. This can be achieved through CRISPR-based reverse genetic screens, where phenotyping is traditionally performed in stable germline knockout (KO) mutants.

Photoconvertible fluorescent proteins: a versatile tool in zebrafish skeletal imaging.

One of the most frequently applied techniques in zebrafish (Danio rerio) research is the visualisation or manipulation of specific cell populations using transgenic reporter lines. The generation of these transgenic zebrafish, displaying cell- or tissue-specific expression of frequently used fluorophores such as Green Fluorescent Protein (GFP) or mCherry, is relatively easy using modern techniques. Fluorophores with different emission wavelengths and driven by different promoters can be monitored simultaneously in the same animal.

Management of Athletes With G6PD Deficiency: Does Missing an Enzyme Mean Missing More Games?

Context: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is likely the most prevalent enzyme deficiency on the planet, with an estimated 4.9% of people, or approximately 330 million individuals, across the globe affected by the disease. In the United States, 4% to 7% of the population is likely affected, but each year our nation's major sport leagues become more international.

CRISPR/Cas9-mediated homology-directed repair by ssODNs in zebrafish induces complex mutational patterns resulting from genomic integration of repair-template fragments.

Targeted genome editing by CRISPR/Cas9 is extremely well fitted to generate gene disruptions, although precise sequence replacement by CRISPR/Cas9-mediated homology-directed repair (HDR) suffers from low efficiency, impeding its use for high-throughput knock-in disease modeling. In this study, we used next-generation sequencing (NGS) analysis to determine the efficiency and reliability of CRISPR/Cas9-mediated HDR using several types of single-stranded oligodeoxynucleotide (ssODN) repair templates for the introduction of disease-relevant point mutations in the zebrafish genome. Our results suggest that HDR rates are strongly determined by repair-template composition, with the most influential factor being homology-arm length.

Zebrafish type I collagen mutants faithfully recapitulate human type I collagenopathies.

The type I collagenopathies are a group of heterogeneous connective tissue disorders, that are caused by mutations in the genes encoding type I collagen and include specific forms of osteogenesis imperfecta (OI) and the Ehlers-Danlos syndrome (EDS). These disorders present with a broad disease spectrum and large clinical variability of which the underlying genetic basis is still poorly understood. In this study, we systematically analyzed skeletal phenotypes in a large set of zebrafish, with diverse mutations in the genes encoding type I collagen, representing different genetic forms of human OI, and a zebrafish model resembling human EDS, which harbors a number of soft connective tissues defects, typical of EDS.

Total ankle arthroplasty versus ankle arthrodesis: a comparative analysis of arc of movement and functional outcomes.

Aims: Few reports compare the contribution of the talonavicular articulation to overall range of movement in the sagittal plane after total ankle arthroplasty (TAA) and tibiotalar arthrodesis. The purpose of this study was to assess changes in ROM and functional outcomes following tibiotalar arthrodesis and TAA.

Patients And Methods: Patients who underwent isolated tibiotalar arthrodesis or TAA with greater than two-year follow-up were enrolled in the study.

The Epidemiology of Fifth Metatarsal Fracture.

Background: A paucity of data exists studying the epidemiology of fifth metatarsal fractures. While a number of studies exist focusing on specific fracture patterns and patient populations, a large comprehensive epidemiologic study on the general public does not.

Objective: We reviewed 1275 fifth metatarsal fractures treated at a multicenter orthopaedic practice attempting to classify mechanism of injury and patient demographics as they pertain to specific fracture patterns.

Outcome of nonoperative management of displaced oblique spiral fractures of the fifth metatarsal shaft.

Background: Nonoperative management has been the preferred treatment for displaced oblique spiral fractures of the fifth metatarsal shaft; yet a paucity of literature supports this claim. The purpose of this investigation was to report the incidence and long-term outcome in the largest cohort of these fractures reported to date.

Methods: From 2006 through 2010, 2990 patients sustaining closed metatarsal fractures were seen and treated.