| Strain Name |
C57BL/6-Tnfrsf1btm1(TNFRSF1B)Bcgen Tnfrsf1atm1(TNFRSF1A)Bcgen Ltbrtm1(LTBR)Bcgen/Bcgen
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Common Name | B-hTNFR2/hTNFR1/hLTBR mice |
| Background | C57BL/6 | Catalog number | 140801 |
| Aliases |
TNFRSF1B, CD120b, TBPII, TNF-R-II, TNF-R75, TNFBR, TNFR1B, TNFR2, TNFR80, p75, p75TNFR, TNF receptor superfamily member 1B; TNFRSF1A, CD120a, FPF, TBP1, TNF-R, TNF-R-I, TNF-R55, TNFAR, TNFR1, TNFR55, TNFR60, p55, p55-R, p60, TNF receptor superfamily member 1A; LTBR, D12S370, LT-BETA-R, TNF-R-III, TNFCR, TNFR-RP, TNFR2-RP, TNFR3, TNFRSF3, lymphotoxin beta receptor |
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Targeting strategy
Gene targeting strategy for B-hTNFR2/hTNFR1/hLTBR mice. The exons 2~6 of mouse Tnfr2 gene that encode the extracellular domain were replaced by human TNFR2 exons 2~6 in B-hTNFR2/hTNFR1/hLTBR mice. The exons 2~6 of mouse Tnfr1 gene that encode the extracellular domain were replaced by human TNFR1 exons 2~6 in B-hTNFR2/hTNFR1/hLTBR mice. The exons 1-7 of mouse Ltbr gene that encode the extracellular domain were replaced by human LTBR exons 1-7 in B-hTNFR2/hTNFR1/hLTBR mice.
Protein expression analysis in regulatory T cells (Tregs)
Strain specific TNFR2 expression analysis in wild-type C57BL/6 mice and homozygous humanized B-hTNFR2/hTNFR1/hLTBR mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTNFR2/hTNFR1/hLTBR mice (H/H; H/H; H/H) stimulated with anti-mouse CD3ε antibody (7.5 μg, i.p.) in vivo for 24 hrs. Protein expression was analyzed with anti-mouse TNFR2 antibody (Biolegend, 113405) and anti-human TNFR2 antibody (BD, 562909) by flow cytometry. Mouse TNFR2 was only detectable in wild-type C57BL/6 mice. Human TNFR2 was exclusively detectable in homozygous B-hTNFR2/hTNFR1/hLTBR mice, but not in wild-type C57BL/6 mice.
Protein expression analysis in granulocytes
Strain specific TNFR1 expression analysis in wild-type C57BL/6 mice and homozygous humanized B-hTNFR2/hTNFR1/hLTBR mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTNFR2/hTNFR1/hLTBR mice (H/H; H/H; H/H). Protein expression was analyzed with anti-mouse TNFR1 antibody (Biolegend, 113005) and anti-human TNFR1 antibody (Biolegend, 369903) by flow cytometry. Mouse TNFR1 was only detectable in wild-type C57BL/6 mice. Human TNFR1 was exclusively detectable in homozygous B-hTNFR2/hTNFR1/hLTBR mice, but not in wild-type C57BL/6 mice.
Protein expression analysis in macrophages
Strain specific LTBR expression analysis in wild-type C57BL/6 mice and homozygous humanized B-hTNFR2/hTNFR1/hLTBR mice by flow cytometry. Spleen, blood and bone marrow were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTNFR2/hTNFR1/hLTBR mice (H/H; H/H; H/H). Protein expression was analyzed with anti-mouse LTBR antibody (Biolegend, 134407) and anti-human LTBR antibody (Biolegend, 322008) by flow cytometry. Mouse LTBR was only detectable in wild-type C57BL/6 mice. Human LTBR was exclusively detectable in homozygous B-hTNFR2/hTNFR1/hLTBR mice, but not in wild-type C57BL/6 mice.
Frequency of leukocyte subpopulations in spleen
Frequency of leukocyte subpopulations in spleen by flow cytometry. Splenocytes were isolated from wild-type C57BL/6 mice and homozygous B-hTNFR2/hTNFR1/hLTBR mice (female, 7-week-old, n=3). A. Flow cytometry analysis of the splenocytes was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. Percentages of T cells, B cells, NK cells, DCs, monocytes, macrophages, neutrophils, CD4+ T cells, CD8+ T cells and Tregs in B-hTNFR2/hTNFR1/hLTBR mice were similar to those in C57BL/6 mice, demonstrating that humanization of TNFR2 and TNFR1 and LTBR do not change the frequency or distribution of these cell types in spleen. Values are expressed as mean ± SEM.
Frequency of leukocyte subpopulations in blood
Frequency of leukocyte subpopulations in blood by flow cytometry. Blood cells were isolated from wild-type C57BL/6 mice and homozygous B-hTNFR2/hTNFR1/hLTBR mice (female, 7-week-old, n=3). A. Flow cytometry analysis of the blood cells was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. Percentages of T cells, B cells, NK cells, DCs, monocytes, macrophages, neutrophils, CD4+ T cells, CD8+ T cells and Tregs in B-hTNFR2/hTNFR1/hLTBR mice were similar to those in C57BL/6 mice, demonstrating that humanization of TNFR2 and TNFR1 and LTBR do not change the frequency or distribution of these cell types in blood. Values are expressed as mean ± SEM.
In vivo efficacy of anti-human LTBR antibody
Antitumor activity of anti-human LTBR antibody (provided by WuXi Biologics) in B-hTNFR2/hTNFR1/hLTBR mice. Murine colon cancer MC38-hLTBR cells (provided by WuXi Biologics) were subcutaneously implanted into homozygous B-hTNFR2/hTNFR1/hLTBR mice (female, 8-9 weeks-old, n=8). Mice were grouped when tumor volume reached approximately 65 mm3, at which time they were intraperitoneally injected with 10mpk anti-human LTBR antibody (provided by WuXi Biologics) indicated in panel. (A) Anti-human LTBR antibody inhibited MC38-hLTBR tumor growth in B-hTNFR2/hTNFR1/hLTBR mice. (B) Body weight changes during treatment. (C) Individual tumor growth curve. As shown in panel A, anti-human LTBR antibody (provided by WuXi Biologics) was efficacious in controlling tumor growth in B-hTNFR2/hTNFR1/hLTBR mice, demonstrating that the B-hTNFR2/hTNFR1/hLTBR mice provide a powerful preclinical model for in vivo evaluation of anti-human LTBR antibody. Values are expressed as mean ± SEM.
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