Limitations of the zebrafish model in hematology research
| Area of research . | Problem . | Possible solution(s) . |
|---|---|---|
| Anatomy | Different morphology of blood cells, eg, erythrocytes and thrombocytes are nucleated | Abstraction in experimental design, ie, seeing similarity rather than difference and learning from difference |
| Different gross anatomy, eg, what is the equivalent of the marrow stroma? | Improving depth of understanding of zebrafish physiology and anatomy will likely narrow rather than widen the apparent gaps | |
| Physiology | Lack of cell markers/antibodies | Partially addressed by rapidly expanding toolbox of fluorescently labeled cell compartments, but generation of a repertoire of zebrafish antibodies is highly desirable |
| Lack of hematopoietic cell lines | Not currently addressed | |
| Lack of biochemical reagents, eg, purified cytokines | Transient factor production based on genetic approaches | |
| Lack of in vitro differentiation system (hematopoietic cell culture assays) | Identification and purification of zebrafish hematopoietic growth factors with optimization of zebrafish cell culture techniques | |
| Lack of inbred strains, eg, for transplantation studies; to facilitate gene mapping | Limited attempts to generate inbred lines which retain hybrid vigor | |
| Genetics | Genetic divergence between fish and humans | Completion of zebrafish genome sequencing project will provide more information about missing genes, but some genetic diseases will prove difficult to model due to genome duplication and divergent evolution |
| Despite divergence in gene structure and function, zebrafish bioassays for human gene activity can still have validity | ||
| No technique for targeted gene modification by homologous recombination | TILLING (Targeting Induced Local Lesions In Genomes) allows ″off the shelf″ ordering of mutants in any gene of interest | |
| Library of characterized insertional mutants; random retroviral insertions disrupt the function of genes and ″tag″ the insertion site within the genome | ||
| Transient knockdown of any gene easily achieved using morpholino antisense oligonucleotides (stable mutant line is preferable because of the possibility of nonspecific/off target morpholino effects) | ||
| Positional cloning challenging while Genome Project still incomplete | Sanger Centre committed to complete Genome Project | |
| Limited options for conditional transgenesis or gene modification | Temperature sensitive alleles | |
| hsp70 (heat shock) promoter | ||
| Gal4-UAS transgenic approaches | ||
| Cre-recombinase or transposon-mediated recombination | ||
| Mutation of fish gene has not always recapitulated the human disease | Consider possibility of quirky phenotypes of individual alleles | |
| Despite some exceptions, fish mutants generally model the human disease persuasively, especially at the level of genetic interactions |
| Area of research . | Problem . | Possible solution(s) . |
|---|---|---|
| Anatomy | Different morphology of blood cells, eg, erythrocytes and thrombocytes are nucleated | Abstraction in experimental design, ie, seeing similarity rather than difference and learning from difference |
| Different gross anatomy, eg, what is the equivalent of the marrow stroma? | Improving depth of understanding of zebrafish physiology and anatomy will likely narrow rather than widen the apparent gaps | |
| Physiology | Lack of cell markers/antibodies | Partially addressed by rapidly expanding toolbox of fluorescently labeled cell compartments, but generation of a repertoire of zebrafish antibodies is highly desirable |
| Lack of hematopoietic cell lines | Not currently addressed | |
| Lack of biochemical reagents, eg, purified cytokines | Transient factor production based on genetic approaches | |
| Lack of in vitro differentiation system (hematopoietic cell culture assays) | Identification and purification of zebrafish hematopoietic growth factors with optimization of zebrafish cell culture techniques | |
| Lack of inbred strains, eg, for transplantation studies; to facilitate gene mapping | Limited attempts to generate inbred lines which retain hybrid vigor | |
| Genetics | Genetic divergence between fish and humans | Completion of zebrafish genome sequencing project will provide more information about missing genes, but some genetic diseases will prove difficult to model due to genome duplication and divergent evolution |
| Despite divergence in gene structure and function, zebrafish bioassays for human gene activity can still have validity | ||
| No technique for targeted gene modification by homologous recombination | TILLING (Targeting Induced Local Lesions In Genomes) allows ″off the shelf″ ordering of mutants in any gene of interest | |
| Library of characterized insertional mutants; random retroviral insertions disrupt the function of genes and ″tag″ the insertion site within the genome | ||
| Transient knockdown of any gene easily achieved using morpholino antisense oligonucleotides (stable mutant line is preferable because of the possibility of nonspecific/off target morpholino effects) | ||
| Positional cloning challenging while Genome Project still incomplete | Sanger Centre committed to complete Genome Project | |
| Limited options for conditional transgenesis or gene modification | Temperature sensitive alleles | |
| hsp70 (heat shock) promoter | ||
| Gal4-UAS transgenic approaches | ||
| Cre-recombinase or transposon-mediated recombination | ||
| Mutation of fish gene has not always recapitulated the human disease | Consider possibility of quirky phenotypes of individual alleles | |
| Despite some exceptions, fish mutants generally model the human disease persuasively, especially at the level of genetic interactions |