B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice

C57BL/6-Il31tm1(IL31)Bcgen Il31ratm1(IL31RA)Bcgen Osmrtm1(OSMR)Bcgen Il4tm2(IL4)Bcgen Il4ratm1(IL4RA)Bcgen/Bcgen • 112908

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B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice

Product nameB-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice
Catalog number112908
Strain nameC57BL/6-Il31tm1(IL31)Bcgen Il31ratm1(IL31RA)Bcgen Osmrtm1(OSMR)Bcgen Il4tm2(IL4)Bcgen Il4ratm1(IL4RA)Bcgen/Bcgen
Strain backgroundC57BL/6
NCBI gene ID133396,3565,3566,386653,9180 (Human)
AliasesBCGF-1; BCGF1; BSF-1; BSF1; IL-4; CD124; IL-4RAA; IL4R; CRL; CRL3; GLM-R; GLMR; GPL; IL-31RA; PLCA2; PRO21384; hGLM-R; IL-31; IL-31R-beta; IL-31RBB; OSMRbeta; PLCA1; OSMR; IL-31; CRL; hGLM-R; IL-31R-beta; OSMR; BCGF-1; IL-4; CD124
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  • Targeting strategy
  • Phenotypic analysis
  • Efficacy
  • Physiological data

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      Targeting strategy

      Gene targeting strategy for B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice.

      • The mouse Il31 gene that encode the full-length protein was replaced by human IL31 counterpart gene sequences in B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice.
      • The coding sequences including human IL31RA extracellular region and mouse Il31ra intracellular region were inserted into mouse IL31RA gene in B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice.
      • The coding sequences including human OSMR extracellular region and mouse Osmr intracellular region were inserted into mouse Osmr gene in B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice.
      • The exons 1-4 of mouse Il4 gene that encode the full length coding sequence were replaced by human IL4 exons 1-4 in B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice.
      • The exons 4-7 of mouse Il4ra gene that encode the extracellular region coding sequences were replaced by human IL4RA exons 4-7 in B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice.

      Note: The mice was obtained by B-hIL31/hIL31RA/hOSMR mice(130746) cross breeding with B-hIL4/hIL4RA mice(120551).

      mRNA expression analysis-IL31

      Strain specific analysis of IL31 mRNA expression in wild-type C57BL/6 mice and B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by RT-PCR. Testis RNA were isolated from wild-type C57BL/6 mice (+/+) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human IL31 primers. Mouse IL31 mRNA was only detectable in wild-type mice. Human IL31 mRNA was exclusively detectable in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice but not in wild-type mice.

      mRNA expression analysis-IL31RA

      Strain specific analysis of IL31RA mRNA expression in wild-type C57BL/6 mice and B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by RT-PCR. Testis RNA were isolated from wild-type C57BL/6 mice (+/+) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human IL31RA primers. Human IL31RA mRNA was exclusively detectable in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice but not in wild-type mice.

      mRNA expression analysis-OSMR

      Strain specific analysis of OSMR mRNA expression in wild-type C57BL/6 mice and B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by RT-PCR. Kidney RNA were isolated from wild-type C57BL/6 mice (+/+) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human OSMR primers. Human OSMR mRNA was exclusively detectable in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice but not in wild-type mice.

      Protein expression analysis

      Strain specific OSMR expression analysis in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by western blot. Brain, liver, lung, and kidney were collected from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (H/H; H/H; H/H; H/H; H/H) and analyzed by western blot with anti-OSMR antibody(Proteintech, 10982-1-AP). m/hOSMR was detectable in brain, liver, lung, and kidney of wild-type C57BL/6JNifdc mice and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice. WT: wild-type C57BL/6JNifdc mice; HO: homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice.

      Strain specific IL4 expression analysis in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by ELISA. Serum were collected from wild-type mice C57BL/6 mice (n=3, 6-week-old, female) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (n=3, 6-week-old, female) that stimulated with anti-mCD3e (7.5 μg/mice, i.p.) and anti-mCD28 (5 μg/mice, i.p.) in vivo for 3 h, and analyzed by ELISA with species-specific IL4 ELISA kit (mIL4 kit: BioLegend 431104; hIL4 kit: BioLegend 430304). Mouse IL4 was only detectable in wild-type mice (A). Human IL4 was exclusively detectable in homozygous mice but not in wild-type mice (B).

      Protein expression analysis in B cells

      Strain specific IL4RA expression analysis in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6JNifdc mice mice and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice stimulated with LPS (200 μg/mice, i.p.) for 2 h, and analyzed by flow cytometry with anti-mouse IL4RA antibody (Biolegend, 144803) or anti-human IL4RA antibody (Biolegend, 355005). Mouse IL4RA was only detectable in wild-type mice, while hIL4RA was exclusively detectable in homozygous mice.

      Frequency of leukocyte subpopulations in spleen

      Frequency of leukocyte subpopulations in spleen by flow cytometry. Splenocytes were isolated from wild-type C57BL/6JNifdc mice (female, 6-week-old, n=3) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (female, 6-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. Frequencies of T cells, B cells, NK cells, DCs, neutrophils, monocytes, macrophages, CD4+ T cells, CD8+ T cells and Tregs in B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice were similar to those in C57BL/6JNifdc mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test. 

      Frequency of leukocyte subpopulations in blood

      Frequency of leukocyte subpopulations in spleen by flow cytometry. Blood cells were isolated from wild-type C57BL/6JNifdc mice (female, 6-week-old, n=3) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (female, 6-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. Frequencies of T cells, B cells, NK cells, DCs, neutrophils, monocytes, macrophages, CD4+ T cells, CD8+ T cells and Tregs in B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice were similar to those in C57BL/6JNifdc mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.

      Frequency of leukocyte subpopulations in lymph nodes

      Frequency of leukocyte subpopulations in spleen by flow cytometry. Lymph nodes cells were isolated from wild-type C57BL/6JNifdc mice (female, 6-week-old, n=3) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (female, 6-week-old, n=3). A. Flow cytometry analysis of the lymph nodes cells was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. Frequencies of T cells, B cells, NK cells, CD4+ T cells, CD8+ T cells and Tregs in B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice were similar to those in C57BL/6JNifdc mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test. 

      Efficacy evaluation of bispecific antibody in AD model of B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA

      Efficacy evaluation of bispecific antibody in AD model of B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice. A. OXA was applied to dorsal and ear skin of B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (female, 8-week-old, n=6)  on day 0, and then challenge to the same site of skin nine times from days 7 to 25. Dupilumab, bispecific antibody (BsAb) (Dupilumab, or BsAb provided by client) were intraperitoneally injected in G3-G7. Ear tissues and serum were collected at the endpoint on day 26. B. Ear thickness. C. Ear thickness change. Scratching behavior was recorded on day 11 (D), day 23 (E). F. Day 16 Serum-Total IgE. G. Day 26 Serum-Total IgE. The results showed that ear thicknesses and serum IgE levels were decreased in TA1,TA2,TA3, and Dupilumab with or without TA4  treated group. The combination of Dupilumab and TA4 significantly attenuates scratching behaviors on day 11. The data is expressed as mean ± SEM (* p< 0.05, ** p< 0.01, *** p< 0.001, **** p< 0.0001). AD: atopic dermatitis; OXA: oxazolone.

      Note: This experiment is a collaboration with the client.

      Efficacy evaluation of bispecific antibody in AD model of B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice. A. Hematoxylin and eosin (H&E) staining. B. Total pathological score. C. Epidermal thickness. D. Eosinophil score. Ear thickness and infiltration scores of eosinophils in ear skin of the groups treated with TA1,TA2 and Dupilumab analog with or without TA4 were decreased significantly compared to that in the isotype control, demonstrating that the B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice provide a powerful preclinical model for in vivo evaluation of bispecific antibody. Infiltration score of eosinophils: 1=slight; 2=mild; 3=moderate; 4=severe. The content of the pathology total score evaluation includes the following aspects: epidermal hyperplasia in skin, erosion/crusting, hyperkeratosis and parakeratosis; inflammatory cell infiltration in dermis and subcutaneous. The data is expressed as mean ± SEM (* p< 0.05, ** p< 0.01, *** p< 0.001, **** p< 0.0001, n.s. no significance).

      Note: This experiment is a collaboration with the client.

      B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice: Growth Curve

      Body weight of wild-type mice and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Wild-type C57BL/6 mice and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice (15 males and 15 females) were monitored for 8-32 weeks to assess overall health of the animals.  Absolute body weight of wild-type mice and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice over time. The weight of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice revealed no abnormalities compared to wild-type controls.

      Hematology analysis-8 weeks

      Complete blood count (CBC) of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Values are expressed as mean ± SD.

      Hematology analysis-16 weeks

      Complete blood count (CBC) of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Values are expressed as mean ± SD.

      Hematology analysis-24 weeks

      Complete blood count (CBC) of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Values are expressed as mean ± SD.

      Hematology analysis-32 weeks

      Complete blood count (CBC) of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Values are expressed as mean ± SD.

      Biochemistry analysis-8 weeks

      Biochemical test of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Values are expressed as mean ± SD.

      Biochemistry analysis-16 weeks

      Biochemical test of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Values are expressed as mean ± SD.

      Biochemistry analysis-24 weeks

      Biochemical test of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Values are expressed as mean ± SD.

      Biochemistry analysis-32 weeks

      Biochemical test of B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. Values are expressed as mean ± SD.

      Gross anatomy of female C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice

      The organs of female C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice (32-week-old, n=15).

      Gross anatomy of male C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice

      The organs of male C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice (32-week-old, n=15).

      Histopathological analysis

      Histopathological analysis of organs in female C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. The main organs of C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice were isolated at 32 weeks of age and analyzed with H&E staining (female, n=3). Results showed that no obvious abnormalities were found in all of the organs (heart, liver, spleen, lung, kidney, brain, stomach, small intestine, large intestine, thymus, lymph nodes, bone marrow, uterus, ovary).

      Histopathological analysis of organs in male C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice. The main organs of C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice were isolated at 32 weeks of age and analyzed with H&E staining (male, n=3). Results showed that no obvious abnormalities were found in all of the organs (heart, liver, spleen, lung, kidney, brain, stomach, small intestine, large intestine, thymus, lymph nodes, bone marrow, testis).

      Organ weight

      Average weight of the main organs of male C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice.

      Average weight of the main organs of male C57BL/6JNifdc and B-hIL31/hIL31/RA/hOSMR/hIL4/hIL4RA mice.

      * When publishing results obtained using this animal model, please acknowledge the source as follows: The animal model [B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice] (Cat# 112908) was purchased from Biocytogen.