Biology subjects

Flik, G.

Publications and source records attributed to Flik, G..

3 recordsLinked to original sources

bioRxiv2020

The transcriptome of regenerating zebrafish scales identifies genes involved in human bone disease

Zebrafish scales are mineralised plates that can regenerate involving de novo bone formation. This presents an opportunity to uncover genes and pathways relevant to human musculoskeletal disease relevant to impaired bone formation. To investigate this hypothesis, we defined transcriptomic profiles of ontogenetic and regenerating scales, and identified 604 differentially expressed genes (DEGs) that were enriched for extracellular matrix, ossification, and cell adhesion pathways. Next, we showed that human orthologues of DEGs were 2.8 times more likely to cause human monogenic skeletal diseases (P<8x10-11), and they showed enrichment for human orthologues associated with polygenetic disease traits including stature, bone density and osteoarthritis (P<0.005). Finally, zebrafish mutants of two human orthologues that were robustly associated with height and osteoarthritis (COL11A2) or bone density only (SPP1) developed skeletal abnormalities consistent with our genetic association studies. Col11a2Y228X/Y228X mutants showed endoskeletal features consistent with abnormal growth and osteoarthritis, whereas spp1P160X/P160X mutants had elevated bone density (P<0.05). In summary, we show that transcriptomic studies of regenerating zebrafish scales have potential to identify new genes and pathways relevant to human skeletal disease.

genetics↗

bioRxiv2019

Early life glucocorticoid exposure modulates immune function in zebrafish (Danio rerio) larvae

In this study we have assessed the effects of increased cortisol levels during early embryonic development on immune function in zebrafish (Danio rerio) larvae. Fertilized eggs were exposed to either a cortisol-containing, a dexamethasone-containing (to stimulate the glucocorticoid receptor selectively) or a control medium for 6 hours post-fertilisation (0-6 hpf). First, we measured baseline expression of a number of immune-related genes (socs3a, mpeg1.1, mpeg1.2 and irg1l) 5 days post-fertilisation (dpf) in larvae of the AB and TL strain to assess the effectiveness of our exposure procedure and potential strain differences. Cortisol and dexamethasone strongly up-regulated baseline expression of these genes independent of strain. The next series of experiments were therefore carried out in larvae of the AB strain only. We measured neutrophil/macrophage recruitment following tail fin amputation (performed at 3 dpf) and phenotypical changes as well as survival following LPS-induced sepsis (150 g/ml; 4-5 dpf). Dexamethasone, but not cortisol, exposure at 0-6 hpf enhanced neutrophil recruitment 4 hours post tail fin amputation. Cortisol and dexamethasone exposure at 0-6 hpf led to a milder phenotype (e.g. less tail fin damage) and enhanced survival following LPS challenge compared to control exposure. Gene-expression analysis showed accompanying differences in transcript abundance of tlr4bb, cxcr4a, myd88, il1{beta} and il10. These data show that early-life exposure to cortisol, which may be considered to be a model or proxy of maternal stress, induces an adaptive response to immune challenges, which seems mediated via the glucocorticoid receptor.

immunology↗

bioRxiv2018

Dissection of intestines from larval zebrafish for molecular analysis

Epigenetic data obtained from whole zebrafish embryos or larvae may mask or dilute organ-specific information. Fluorescence activated cell sorting can diverge cells from their native state, and cryosections often yield insufficient material for molecular analysis. Here, we present a reproducible method for larval intestinal isolation at 5, 7, and 9 days post-fertilization, using the intestine-specific transgene tgBAC(cldn15la:GFP). With tweezers, the intestine can be pulled out of the abdomen in one smooth motion. Upon removal of adhering tissues, intestines can be directly used for analyses. Each dissection takes 3-6 minutes per fish. We demonstrate that 10 and 25 dissected intestines yield enough material for RNA-sequencing and ChIP-sequencing, respectively. This method results in high quality, live material, suitable for many downstream applications.\n\nMETHOD SUMMARYWe present a reproducible method for zebrafish larval intestinal isolation which results in high quality, live material. With tweezers, the intestine can be pulled out of the abdomen and after removal of adhering tissues, intestines can be directly used for analyses. We demonstrate that 10 and 25 dissected intestines yield enough material for RNA-sequencing and ChIP-sequencing, respectively.

developmental biology↗