B6;129S-Sell^tm3Hyn Selp^tm1Hyn/J

Stock number: 003805
Product name: PL-
Research areas: Immunology, Inflammation and Autoimmunity Research

Description

These double knock-out mice exhibit reductions in leukocyte rolling and in extravasation of neutrophils in thioglycollate-induced peritonitis.

Detailed description

Mice that are homozygous null for the Sell and Selp genes are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. No Sell and Selp gene products (mRNA or protein) are detected. Leukocytes from these mice exhibit a deficiency in the ability to interact with, and roll along, the venular wall endothelium. Such a deficiency in the crucial first step of leukocyte recruitment to surrounding tissues in response to infection or injury results in elevated leukocyte counts in the peripheral blood. These defects are largely due to absence of Selp. In addition, lack of Sell leads to failure of lymphocytes to enter lymph nodes and gthus to hypoplastic lymph nodes. These mice are suitable for use in research applications studying leukocyte homing, infectious diseases and inflammation.

Development

Sequential mutation by homologous recombination was utilized to replace the Sell and Selp genes with drug-resistance genes for neomycin and puromycin (respectively) in 129S2/SvPas derived D3 embryonic stem (ES) cells. This approach was necessary because the Sell and Selp genes are too closely linked for double-deficient animals to be generated by mating single-deficient animals. Correctly targeted ES cells were injected into C57BL/6 blastocysts and chimeric offspring obtained.

Allele

Symbol: Sell^tm3Hyn
Name: targeted mutation 3, Richard Hynes
MGI accession ID: MGI:3839977
Generation method: Targeted
Attributes: Null/Knockout
Marker symbol: Sell
Marker name: selectin, lymphocyte
Chromosome: 1
Strain of origin: 129S2/SvPas
Molecular note: The three selectin genes, Sele, Sell, and Selp, are found within a 300 kb genomic region. To generate double knockouts, ES cells heterozygote for the Selp^tm1Hyn and Sele^tm2Hyn null alleles in trans to each other were targeted a third time. A PGK-puro cassette was used to delete a 2.1 kb region containing the exons encoding the lectin domain and most of the epidermal growth factor domain. Flow cytometric analysis of homozygote leukocytes showed an absence of the L-selectin and P-selectin protein while E-selectin expression was unaffected. Thus, this targeted mutation occurred in cis with the Selp^tm1Hyn allele and in trans with the Sele^tm2Hyn allele. Due to their genomic proximity, the Sell^tm3Hyn and Selp^tm1Hyn alleles should propagate together through the offspring.

Allele

Symbol: Selp^tm1Hyn
Name: targeted mutation 1, Richard Hynes
MGI accession ID: MGI:1857245
Generation method: Targeted
Attributes: Null/Knockout
Marker symbol: Selp
Marker name: selectin, platelet
Chromosome: 1
Strain of origin: 129S2/SvPas
Molecular note: A portion of exon 3 encoding 10 amino acids of a signal peptide and 27 amino acids of the lectin domain was replaced with a PGK-neo cassette. Mice were treated with lipopolysaccharide to augment the normally low levels of selectin in lung and liver tissue. A longer transcript, resulting from aberrant or cryptic splicing involving the neomycin cassette, was detected at low levels in mutant mice by Northern blot analysis. RT-PCR further confirmed the presence of low levels of transcript in homozygous mice. Immunofluorescence analysis of activated platelets and lung sections from homozygous mutant mice showed an absence of encoded protein. These results were further confirmed by flow cytometric analysis of activated platelets and metabolic labeling of lung tissue.