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Zebrafish (Danio rerio) are aquatic vertebrates with significant homology to their terrestrial counterparts. While zebrafish have a centuries-long track record in developmental and regenerative biology, their utility has grown exponentially with the onset of modern genetics. This is exemplified in studies focused on skeletal development and repair. Herein, the numerous contributions of zebrafish to our understanding of the basic science of cartilage, bone, tendon/ligament, and other skeletal tissues are described, with a particular focus on applications to development and regeneration. We summarize the genetic strengths that have made the zebrafish a powerful model to understand skeletal biology. We also highlight the large body of existing tools and techniques available to understand skeletal development and repair in the zebrafish and introduce emerging methods that will aid in novel discoveries in skeletal biology. Finally, we review the unique contributions of zebrafish to our understanding of regeneration and highlight diverse routes of repair in different contexts of injury. We conclude that zebrafish will continue to fill a niche of increasing breadth and depth in the study of basic cellular mechanisms of skeletal biology.

PurposeThe exocyst complex is a conserved protein complex that mediates fusion of intracellular vesicles to the plasma membrane and is implicated in processes including cell polarity, cell migration, ciliogenesis, cytokinesis, autophagy, and fusion of secretory vesicles. The essential role of these genes in human genetic disorders, however, is unknown.MethodsWe performed homozygosity mapping and exome sequencing of consanguineous families with recessively inherited brain development disorders. We modeled an EXOC7 splice variant in vitro and examined EXOC7 messenger RNA (mRNA) expression in developing mouse and human cortex. We modeled exoc7 loss-of-function in a zebrafish knockout.ResultsWe report variants in exocyst complex members, EXOC7 and EXOC8, in a novel disorder of cerebral cortex development. In EXOC7, we identified four independent partial loss-of-function (LOF) variants in a recessively inherited disorder characterized by brain atrophy, seizures, and developmental delay, and in severe cases, microcephaly and infantile death. In EXOC8, we found a homozygous truncating variant in a family with a similar clinical disorder. We modeled exoc7 deficiency in zebrafish and found the absence of exoc7 causes microcephaly.ConclusionOur results highlight the essential role of the exocyst pathway in normal cortical development and how its perturbation causes complex brain disorders.

Intervertebral disc disease (IDD) is a debilitating spine condition that can be caused by intervertebral disc (IVD) damage which progresses towards IVD degeneration and dysfunction. Recently, human pluripotent stem cells (hPSCs) were recognized as a valuable resource for cell-based regenerative medicine in skeletal diseases. Therefore, adult somatic cells reprogrammed into human induced pluripotent stem cells (hiPSCs) represent an attractive cell source for the derivation of notochordal-like cells (NCs) as a first step towards the development of a regenerative therapy for IDD. Utilizing a differentiation method involving treatment with a four-factor cocktail targeting the BMP, FGF, retinoic acid, and Wnt signaling pathways, we differentiate CRISPR/Cas9-generated mCherry-reporter knock-in hiPSCs into notochordal-like cells. Comprehensive analysis of transcriptomic changes throughout the differentiation process identified regulation of histone methylation as a pivotal driver facilitating the differentiation of hiPSCs into notochordal-like cells. We further provide evidence that specific inhibition of histone demethylases KDM2A and KDM7A/B enhanced the lineage commitment of hiPSCs towards notochordal-like cells. Our results suggest that inhibition of KDMs could be leveraged to alter the epigenetic landscape of hiPSCs to control notochord-specific gene expression. Thus, our study highlights the importance of epigenetic regulators in stem cell-based regenerative approaches for the treatment of disc degeneration. [ABSTRACT FROM AUTHOR]

Katanin is a microtubule-severing complex whose catalytic activities are well characterized, but whose in vivo functions are incompletely understood. Human mutations in KATNB1, which encodes the noncatalytic regulatory p80 subunit of katanin, cause severe microlissencephaly. Loss of Katnb1 in mice confirms essential roles in neurogenesis and cell survival, while loss of zebrafish katnb1 reveals specific roles for katnin p80 in early and late developmental stages. Surprisingly, Katnb1 null mutant mouse embryos display hallmarks of aberrant Sonic hedgehog signaling, including holoprosencephaly. KATNB1-deficient human cells show defective proliferation and spindle structure, while Katnb1 null fibroblasts also demonstrate a remarkable excess of centrioles, with supernumerary cilia but deficient Hedgehog signaling. Our results reveal unexpected functions for KATNB1 in regulating overall centriole, mother centriole, and cilia number, and as an essential gene for normal Hedgehog signaling during neocortical development.

Subacute thyroiditis (SAT) is a self-limited inflammatory disease and a rare cause of thyrotoxicosis. Although the exact etiology of SAT is not sufficiently understood, it is generally associated to viral infections. Current evidence highlights that SAT may be a potentially uncommon manifestation of ongoing Coronavirus disease 2019 (COVID-19) infection or a post-viral complication of the disease. Despite that SAT is a rare manifestation associated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) disease both in ongoing and resolved COVID-19 infection, the ever-increasing numbers of COVID-19 patients strengthens the possibility that this particular disease entity will be of more immediate concern in the future. The current work aims to summarize the approach of SARS-CoV-2-associated SAT, present its pathophysiology, outline current research evidence found in the literature, and discuss potential differential diagnoses and diagnostic dilemmas through an illustrative case. [ABSTRACT FROM AUTHOR]

Germ cell differentiation and maintenance relies on complex regulation of mitotic and meiotic progression. Cyclin-dependent kinases (CDKs) and their activating cyclin partners are known to have specialized roles in regulating cell cycle progression across tissues, including germ cells. Very little is known about CDK/cyclin function in zebrafish or the regulation of germ cell maintenance and differentiation. In a forward genetic screen for gonadogenesis defects in zebrafish, a mutation disrupting cdk21 (cyclin-dependent kinase 21) was identified, which caused gonad hypoplasia, reduced fertility and failure of female sex specification. The cdk21 gene is unique to fishes, though the encoded protein is related to the D-cyclin partners Cdk4 and Cdk6, which are known G1 cell cycle regulators. In the testis, cdk21 mutant germ cells exhibited cell cycle defects such as diminished proliferation, prolonged meiosis and delayed sperm differentiation. Furthermore, cdk21 mutants failed to maintain germ cells following breeding. Based on these findings, we propose that cdk21 regulates spermatogonial proliferation, progression through meiosis and germline stem cell activation in the testis. In addition, we investigated cdk4 and cdk6 in zebrafish development and found that each has distinct expression patterns in the gonads. Mutant analysis demonstrated that cdk6 was necessary for viability beyond larval stages. In contrast, cdk4 mutants were viable but were all male with low breeding success and sperm overabundance. Our analysis demonstrated that zebrafish harbor three genes of the cdk4/6 family, cdk4, cdk6 and cdk21, with cdk21 having an essential role in germ cell development in the testis.