scholarly journals Zebrafish model for spondylo-megaepiphyseal-metaphyseal dysplasia reveals post-embryonic roles of Nkx3.2 in the skeleton

Development ◽  
2021 ◽  
Vol 148 (2) ◽  
pp. dev193409
Author(s):  
Joanna Smeeton ◽  
Natasha Natarajan ◽  
Arati Naveen Kumar ◽  
Tetsuto Miyashita ◽  
Pranidhi Baddam ◽  
...  
Keyword(s):  
Skeletal Development ◽  
Facial Skeleton ◽  
Zebrafish Model ◽  
Growth Plates ◽  
Metaphyseal Dysplasia ◽  
Embryonic Lethal ◽  
Skeletal Defects ◽  
Single Cell Rna Sequencing ◽  
Jaw Joint

ABSTRACTThe regulated expansion of chondrocytes within growth plates and joints ensures proper skeletal development through adulthood. Mutations in the transcription factor NKX3.2 underlie spondylo-megaepiphyseal-metaphyseal dysplasia (SMMD), which is characterized by skeletal defects including scoliosis, large epiphyses, wide growth plates and supernumerary distal limb joints. Whereas nkx3.2 knockdown zebrafish and mouse Nkx3.2 mutants display embryonic lethal jaw joint fusions and skeletal reductions, respectively, they lack the skeletal overgrowth seen in SMMD patients. Here, we report adult viable nkx3.2 mutant zebrafish displaying cartilage overgrowth in place of a missing jaw joint, as well as severe dysmorphologies of the facial skeleton, skullcap and spine. In contrast, cartilage overgrowth and scoliosis are absent in rare viable nkx3.2 knockdown animals that lack jaw joints, supporting post-embryonic roles for Nkx3.2. Single-cell RNA-sequencing and in vivo validation reveal increased proliferation and upregulation of stress-induced pathways, including prostaglandin synthases, in mutant chondrocytes. By generating a zebrafish model for the skeletal overgrowth defects of SMMD, we reveal post-embryonic roles for Nkx3.2 in dampening proliferation and buffering the stress response in joint-associated chondrocytes.

The type I BMP receptor BMPRIB is required for chondrogenesis in the mouse limb

Development ◽  
2000 ◽  
Vol 127 (3) ◽  
pp. 621-630 ◽  
Author(s):  
S.E. Yi ◽  
A. Daluiski ◽  
R. Pederson ◽  
V. Rosen ◽  
K.M. Lyons
Keyword(s):  
Skeletal Development ◽  
Embryonic Stem ◽  
Appendicular Skeleton ◽  
Type I ◽  
Targeted Disruption ◽  
Unique Role ◽  
Bmp Receptors ◽  
Skeletal Defects ◽  
Mouse Limb

Mice carrying a targeted disruption of BmprIB were generated by homologous recombination in embryonic stem cells. BmprIB(−/−) mice are viable and, in spite of the widespread expression of BMPRIB throughout the developing skeleton, exhibit defects that are largely restricted to the appendicular skeleton. Using molecular markers, we show that the initial formation of the digital rays occurs normally in null mutants, but proliferation of prechondrogenic cells and chondrocyte differentiation in the phalangeal region are markedly reduced. Our results suggest that BMPRIB-mediated signaling is required for cell proliferation after commitment to the chondrogenic lineage. Analyses of BmprIB and Gdf5 single mutants, as well as BmprIB; Gdf5 double mutants suggests that GDF5 is a ligand for BMPRIB in vivo. BmprIB; Bmp7 double mutants were constructed in order to examine whether BMPRIB has overlapping functions with other type I BMP receptors. BmprIB; Bmp7 double mutants exhibit severe appendicular skeletal defects, suggesting that BMPRIB and BMP7 act in distinct, but overlapping pathways. These results also demonstrate that in the absence of BMPRIB, BMP7 plays an essential role in appendicular skeletal development. Therefore, rather than having a unique role, BMPRIB has broadly overlapping functions with other BMP receptors during skeletal development.

scholarly journals Skeletal defects in VEGF120/120 mice reveal multiple roles for VEGF in skeletogenesis

Development ◽  
2002 ◽  
Vol 129 (8) ◽  
pp. 1893-1904 ◽  
Author(s):  
Elazar Zelzer ◽  
William McLean ◽  
Yin-Shan Ng ◽  
Naomi Fukai ◽  
Anthony M. Reginato ◽  
...  
Keyword(s):  
Blood Vessels ◽  
Skeletal Development ◽  
Surrounding Tissue ◽  
Ossification Center ◽  
Direct Role ◽  
Osteoblastic Activity ◽  
Chondrocyte Maturation ◽  
Skeletal Defects

Angiogenesis is an essential component of skeletal development and VEGF signaling plays an important if not pivotal role in this process. Previous attempts to examine the roles of VEGF in vivo have been largely unsuccessful because deletion of even one VEGF allele leads to embryonic lethality before skeletal development is initiated. The availability of mice expressing only the VEGF120 isoform (which do survive to term) has offered an opportunity to explore the function of VEGF during embryonic skeletal development. Our study of these mice provides new in vivo evidence for multiple important roles of VEGF in both endochondral and intramembranous bone formation, as well as some insights into isoform-specific functions. There are two key differences in vascularization of developing bones between wild-type and VEGF120/120 mice. VEGF120/120 mice have not only a delayed recruitment of blood vessels into the perichondrium but also show delayed invasion of vessels into the primary ossification center, demonstrating a significant role of VEGF at both an early and late stage of cartilage vascularization. These findings are the basis for a two-step model of VEGF-controlled vascularization of the developing skeleton, a hypothesis that is supported by the new finding that VEGF is expressed robustly in the perichondrium and surrounding tissue of cartilage templates of future bones well before blood vessels appear in these regions. We also describe new in vivo evidence for a possible role of VEGF in chondrocyte maturation, and document that VEGF has a direct role in regulating osteoblastic activity based on in vivo evidence and organ culture experiments.

scholarly journals Overexpression of Notch Signaling Induces Hyperosteogeny in Zebrafish

2019 ◽  
Vol 20 (15) ◽  
pp. 3613 ◽  
Author(s):  
Sung-Tzu Liang ◽  
Jung-Ren Chen ◽  
Jhih-Jie Tsai ◽  
Yu-Heng Lai ◽  
Chung-Der Hsiao
Keyword(s):  
Notch Signaling ◽  
Skeletal Development ◽  
Transgenic Zebrafish ◽  
Bone Homeostasis ◽  
Zebrafish Model ◽  
Therapeutic Drugs ◽  
Evolutionarily Conserved ◽  
Multicellular Organisms

Notch signaling is one of the evolutionarily conserved signaling pathways in multicellular organisms. It plays an important role in embryonic development. During skeletal development of vertebrates, it regulates bone homeostasis by manipulating both osteoblastogenesis and osteoclastogenesis through different mechanisms. However, due to the different nature of Notch signaling in mesenchymal stem cell and osteoblast, regulation of Notch signaling in bone-related diseases remains unsettled. Previous studies by cell culture and mouse models showed contradictory results regarding the role of Notch signaling in bone homeostasis. To clarify the role of Notch signaling in osteogenesis, we established a zebrafish model, in which Notch1a intracellular domain (N1aICD) was specifically expressed in the osteoblasts. We found that overexpression of N1aICD in osteoblasts caused hyperosteogeny in the column region of zebrafish with the morphology of narrowed neural/hemal canals. Moreover, increased metabolic activity of osteoblasts instead of augmenting osteoblast number led to hyperosteogeny in N1aICD-overexpressed zebrafish. In summary, we successfully established a transgenic zebrafish line overexpressing N1aICD to clarify the in-vivo function of Notch signaling during osteoblastogenesis. In the future, this fish line can serve as a valuable tool to test the therapeutic drugs for hyperosteogeny.

scholarly journals Comprehensive series of Irx cluster mutants reveals diverse roles in facial cartilage development

Development ◽  
2021 ◽  
Author(s):  
D'Juan T. Farmer ◽  
Punam Patel ◽  
Rachelle Choi ◽  
Chih-Yu Liu ◽  
J. Gage Crump
Keyword(s):  
Function Analysis ◽  
Skeletal Development ◽  
Proper Function ◽  
Loss Of Function ◽  
Facial Skeleton ◽  
Cartilage Development ◽  
The Face ◽  
Vertebrate Skeleton ◽  
And Cartilage

Proper function of the vertebrate skeleton requires the development of distinct articulating embryonic cartilages. Irx transcription factors are arranged in co-regulated clusters that are expressed in the developing skeletons of the face and appendages. IrxB cluster genes are required for the separation of toes in mice and formation of the hyoid joint in zebrafish, yet whether Irx genes had broader roles in skeletal development remained unclear. Here we perform a comprehensive loss-of-function analysis of all 11 Irx genes in zebrafish. We uncover conserved requirements for IrxB genes in formation of the fish and mouse scapula. In the face, we find a requirement for IrxAb genes and irx7 in formation of anterior neural crest precursors of the jaw, and for IrxBa genes in formation of endodermal pouches and gill cartilages. We also observe extensive joint loss and cartilage fusions in animals with combinatorial losses of Irx clusters, with in vivo imaging revealing that at least some of these fusions arise through inappropriate chondrogenesis. Our analysis reveals diverse roles for Irx genes in the formation and later segmentation of the facial skeleton.

scholarly journals Single-cell RNA sequencing reveals ex vivo signatures of SARS-CoV-2-reactive T cells through ‘reverse phenotyping’

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
David S. Fischer ◽  
Meshal Ansari ◽  
Karolin I. Wagner ◽  
Sebastian Jarosch ◽  
Yiqi Huang ◽  
...  
Keyword(s):  
T Cells ◽  
Single Cell ◽  
Rna Sequencing ◽  
Ex Vivo ◽  
Severe Disease ◽  
Human Leukocyte ◽  
Cell Receptor ◽  
Single Cell Rna Sequencing

AbstractThe in vivo phenotypic profile of T cells reactive to severe acute respiratory syndrome (SARS)-CoV-2 antigens remains poorly understood. Conventional methods to detect antigen-reactive T cells require in vitro antigenic re-stimulation or highly individualized peptide-human leukocyte antigen (pHLA) multimers. Here, we use single-cell RNA sequencing to identify and profile SARS-CoV-2-reactive T cells from Coronavirus Disease 2019 (COVID-19) patients. To do so, we induce transcriptional shifts by antigenic stimulation in vitro and take advantage of natural T cell receptor (TCR) sequences of clonally expanded T cells as barcodes for ‘reverse phenotyping’. This allows identification of SARS-CoV-2-reactive TCRs and reveals phenotypic effects introduced by antigen-specific stimulation. We characterize transcriptional signatures of currently and previously activated SARS-CoV-2-reactive T cells, and show correspondence with phenotypes of T cells from the respiratory tract of patients with severe disease in the presence or absence of virus in independent cohorts. Reverse phenotyping is a powerful tool to provide an integrated insight into cellular states of SARS-CoV-2-reactive T cells across tissues and activation states.

scholarly journals 5-Bromoprotocatechualdehyde Combats against Palmitate Toxicity by Inhibiting Parkin Degradation and Reducing ROS-Induced Mitochondrial Damage in Pancreatic β-Cells

Antioxidants ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 264
Author(s):  
Seon-Heui Cha ◽  
Chunying Zhang ◽  
Soo-Jin Heo ◽  
Hee-Sook Jun
Keyword(s):  
Cell Loss ◽  
Cellular Damage ◽  
Mitochondrial Morphology ◽  
Protective Effects ◽  
Β Cells ◽  
Β Cell ◽  
Zebrafish Model ◽  
Cell Dysfunction ◽  
Glucose Stimulated Insulin Secretion

Pancreatic β-cell loss is critical in diabetes pathogenesis. Up to now, no effective treatment has become available for β-cell loss. A polyphenol recently isolated from Polysiphonia japonica, 5-Bromoprotocatechualdehyde (BPCA), is considered as a potential compound for the protection of β-cells. In this study, we examined palmitate (PA)-induced lipotoxicity in Ins-1 cells to test the protective effects of BPCA on insulin-secreting β-cells. Our results demonstrated that BPCA can protect β-cells from PA-induced lipotoxicity by reducing cellular damage, preventing reactive oxygen species (ROS) overproduction, and enhancing glucose-stimulated insulin secretion (GSIS). BPCA also improved mitochondrial morphology by preserving parkin protein expression. Moreover, BPCA exhibited a protective effect against PA-induced β-cell dysfunction in vivo in a zebrafish model. Our results provide strong evidence that BPCA could be a potential therapeutic agent for the management of diabetes.

scholarly journals Loss of Wnt16 Leads to Skeletal Deformities and Downregulation of Bone Developmental Pathway in Zebrafish

2021 ◽  
Vol 22 (13) ◽  
pp. 6673
Author(s):  
Xiaochao Qu ◽  
Mei Liao ◽  
Weiwei Liu ◽  
Yisheng Cai ◽  
Qiaorong Yi ◽  
...  
Keyword(s):  
Gene Knockout ◽  
Integration Site ◽  
Skeletal Development ◽  
Rna Seq ◽  
Developmental Pathway ◽  
Mineral Density ◽  
Zebrafish Model ◽  
Protein Protein Interaction ◽  
Ct Analysis

Wingless-type MMTV integration site family, member 16 (wnt16), is a wnt ligand that participates in the regulation of vertebrate skeletal development. Studies have shown that wnt16 can regulate bone metabolism, but its molecular mechanism remains largely undefined. We obtained the wnt16-/- zebrafish model using the CRISPR-Cas9-mediated gene knockout screen with 11 bp deletion in wnt16, which led to the premature termination of amino acid translation and significantly reduced wnt16 expression, thus obtaining the wnt16-/- zebrafish model. The expression of wnt16 in bone-related parts was detected via in situ hybridization. The head, spine, and tail exhibited significant deformities, and the bone mineral density and trabecular bone decreased in wnt16-/- using light microscopy and micro-CT analysis. RNA sequencing was performed to explore the differentially expressed genes (DEGs). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis found that the down-regulated DEGs are mainly concentrated in mTOR, FoxO, and VEGF pathways. Protein–protein interaction (PPI) network analysis was performed with the detected DEGs. Eight down-regulated DEGs including akt1, bnip4, ptena, vegfaa, twsg1b, prkab1a, prkab1b, and pla2g4f.2 were validated by qRT-PCR and the results were consistent with the RNA-seq data. Overall, our work provides key insights into the influence of wnt16 gene on skeletal development.

scholarly journals Development of BCR-ABL1 Transgenic Zebrafish Model Reproducing Chronic Myeloid Leukemia (CML) Like-Disease and Providing a New Insight into CML Mechanisms

Cells ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 445
Author(s):  
Daniela Zizioli ◽  
Simona Bernardi ◽  
Marco Varinelli ◽  
Mirko Farina ◽  
Luca Mignani ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Fusion Gene ◽  
Philadelphia Chromosome ◽  
Leukemic Cells ◽  
Transgenic Zebrafish ◽  
Hematopoietic Tissue ◽  
Zebrafish Model ◽  
Altered Expression

Zebrafish has proven to be a versatile and reliable experimental in vivo tool to study human hematopoiesis and model hematological malignancies. Transgenic technologies enable the generation of specific leukemia types by the expression of human oncogenes under specific promoters. Using this technology, a variety of myeloid and lymphoid malignancies zebrafish models have been described. Chronic myeloid leukemia (CML) is a clonal myeloproliferative neoplasia characterized by the BCR-ABL1 fusion gene, derived from the t (9;22) translocation causing the Philadelphia Chromosome (Ph). The BCR-ABL1 protein is a constitutively activated tyrosine kinas inducing the leukemogenesis and resulting in an accumulation of immature leukemic cells into bone marrow and peripheral blood. To model Ph+ CML, a transgenic zebrafish line expressing the human BCR-ABL1 was generated by the Gal4/UAS system, and then crossed with the hsp70-Gal4 transgenic line. The new line named (BCR-ABL1pUAS:CFP/hsp70-Gal4), presented altered expression of hematopoietic markers during embryonic development compared to controls and transgenic larvae showed proliferating hematopoietic cells in the caudal hematopoietic tissue (CHT). The present transgenic zebrafish would be a robust CML model and a high-throughput drug screening tool.

scholarly journals Anti-seizure activity of African medicinal plants: the identification of bioactive alkaloids from the stem bark of Rauvolfia caffra using an in vivo zebrafish model

2021 ◽  
pp. 114282
Author(s):  
Talent Chipiti ◽  
Alvaro M. Viljoen ◽  
Maria L. Cordero-Maldonado ◽  
Clinton.G.L. Veale ◽  
Fanie R. Van Heerden ◽  
...  
Keyword(s):  
Medicinal Plants ◽  
Seizure Activity ◽  
Stem Bark ◽  
Zebrafish Model

In vivo SAR and STR analyses of alkaloids from Picrasma quassioides identify 1-hydroxymethyl-8-hydroxy-β-carboline as a novel natural angiogenesis inhibitor

RSC Advances ◽  
2016 ◽  
Vol 6 (12) ◽  
pp. 9484-9494 ◽  
Author(s):  
Guiyi Gong ◽  
Qinghua Lin ◽  
Jian Xu ◽  
Feng Ye ◽  
Lingling Jiang ◽  
...  
Keyword(s):  
Angiogenesis Inhibitor ◽  
Zebrafish Model ◽  
Picrasma Quassioides

Twenty alkaloids were obtained from the anti-angiogenic fraction of Picrasma quassioides and their SAR/STR were studies by a zebrafish model. We had identified 3 as an angiogenesis inhibitor and confirmed in vitro.

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