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Strain name
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C57BL/6-Tslptm1(TSLP)Crlf2tm2(CRLF2)/Bcgen
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Common Name
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B-hTSLP/hTSLPR mice plus
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Background
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C57BL/6
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Catalog number
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112744
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Aliases
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TSLP: NA;CRLF2: CRL2Y, TSLPR
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Protein expression analysis-TSLP
Strain specific TSLP expression analysis in wild type C57BL/6 mice, homozygous B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus by ELISA. Calcipotriol (MC903) was dissolved in ethanol and topically applied on ears of wild type C57BL/6 mice, homozygous B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus for 7 days. Ear grinding supernatant from the three strain of mice were analyzed by ELISA. Mouse TSLP was only detectable in wild type C57BL/6 mice. Human TSLP was detectable in homozygous B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus but not in wild type mice.
Protein expression analysis of TSLPR in spleen
Data from Huabo Biopharm Co. Ltd.
Mouse and human TSLPR expression analysis in splenocytes. Splenocytes were collected from wild type C57BL/6 mice, homozygous B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus. TSLPR expressed on cDC, pDC and non DCs were analyzed by flow cytometry with species-specific anti-TSLPR antibody. Mouse TSLPR were detectable on cDC, pDC and non DCs of wild type C57BL/6 mice, but not on the cells of TSLPR humanized mice. Human TSLPR was detectable on cDC, pDC and non DCs of B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus, but not on the cells of wild type C57BL/6 mice.
Protein expression analysis of TSLPR in bone marrow
Data from Huabo Biopharm Co. Ltd.
Mouse and human TSLPR expression analysis in bone marrow. Bone marrow cells were collected from wild type C57BL/6 mice, homozygous B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus. TSLPR expressed on bone marrow cells were analyzed by flow cytometry with species-specific anti-TSLPR antibody. Mouse TSLPR were detectable on cDC, pDC and non DCs in wild type C57BL/6 mice, but not on the cells of TSLPR humanized mice. Human TSLPR was detectable on cDC, pDC and non DCs of B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus, but not on the cells of wild type C57BL/6 mice.
Protein expression analysis of hTSLPR in cDC1 from bone marrow
Data from Huabo Biopharm Co. Ltd.
Human TSLPR expression analysis in cDC1 from bone marrow. Bone marrow were collected from wild type C57BL/6 mice, homozygous B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus. Human TSLPR expressed on cDC1 were analyzed by flow cytometry with species-specific antibodies. Human TSLPR was highly expressed on the cDC1 in B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus.
Analysis of leukocytes cell subpopulation in spleen
Analysis of spleen leukocyte subpopulations by flow cytometry. Splenocytes were isolated from female C57BL/6 and B-hTSLP/hTSLPR mice plus (n=3, 9-week-old). Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live cells were gated for the CD45+ population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of T cells, B cells, NK cells, DCs, neutrophils, monocytes and macrophages in homozygous B-hTSLP/hTSLPR mice plus were similar to those in the C57BL/6 mice, demonstrating that TSLP，TSLPR and IL7R humanized does not change the overall development, differentiation or distribution of these cell types in splenocytes. Values are expressed as mean ± SEM.
Analysis of T cell subpopulation in spleen
Analysis of spleen T cell subpopulations by flow cytometry. Splenocytes were isolated from female C57BL/6 and B-hTSLP/hTSLPR mice plus (n=3, 9-week-old). Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live CD45+ cells were gated for CD3+ T cell population and used for further analysis as indicated here. B. Results of FACS analysis. The percent of CD4+ T cells, CD8+ T cells and Tregs in homozygous B-hTSLP/hTSLPR mice plus were similar to those in the C57BL/6 mice, demonstrating that TSLP，TSLPR and IL7R humanized does not change the overall development, differentiation or distribution of these T cell types in spleen. Values are expressed as mean ± SEM.
Analysis of leukocytes cell subpopulation in blood
Analysis of blood leukocyte subpopulations by flow cytometry. Blood were isolated from female C57BL/6 and B-hTSLP/hTSLPR mice plus (n=3, 9-week-old). Flow cytometry analysis of the blood was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live cells were gated for the CD45+ population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of T cells, B cells, NK cells, dendritic cells, neutrophils, monocytes and macrophages in homozygous B-hTSLP/hTSLPR mice plus were similar to those in the C57BL/6 mice, demonstrating that TSLP，TSLPR and IL7R humanized does not change the overall development, differentiation or distribution of these cell types in blood. Values are expressed as mean ± SEM.
Analysis of T cell subpopulation in blood
Analysis of blood T cell subpopulations by flow cytometry. Blood were isolated from female C57BL/6 and B-hTSLP/hTSLPR mice plus (n=3, 9-week-old). Flow cytometry analysis of the blood was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live CD45+ cells were gated for CD3+ T cell population and used for further analysis as indicated here. B. Results of FACS analysis. The percent of CD4+ T cells, CD8+ T cells and Tregs in homozygous B-hTSLP/hTSLPR mice plus were similar to those in the C57BL/6 mice, demonstrating that TSLP，TSLPR and IL7R humanized does not change the overall development, differentiation or distribution of these T cell types in blood. Values are expressed as mean ± SEM.
Analysis of leukocytes cell subpopulation in lymph node
Analysis of lymph node leukocyte subpopulations by flow cytometry. Lymph nodes were isolated from female C57BL/6 and B-hTSLP/hTSLPR mice plus (n=3, 9-week-old). Flow cytometry analysis of the lymph node was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live cells were gated for the CD45+ population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of T cells, B cells, NK cells, dendritic cells, neutrophils, monocytes and macrophages in homozygous B-hTSLP/hTSLPR mice plus were similar to those in the C57BL/6 mice, demonstrating that TSLP, TSLPR and IL7R humanized does not change the overall development, differentiation or distribution of these cell types in lymph node. Values are expressed as mean ± SEM.
Analysis of T cell subpopulation in lymph node
Analysis of lymph node T cell subpopulations by flow cytometry. Lymph nodes were isolated from female C57BL/6 and B-hTSLP/hTSLPR mice plus (n=3, 9-week-old). Flow cytometry analysis of the lymph node was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live CD45+ cells were gated for CD3+ T cell population and used for further analysis as indicated here. B. Results of FACS analysis. The percent of CD4+ T cells, CD8+ T cells and Tregs in homozygous B-hTSLP/hTSLPR mice plus were similar to those in the C57BL/6 mice, demonstrating that TSLP, TSLPR and IL7R humanized does not change the overall development, differentiation or distribution of these T cell types in lymph node. Values are expressed as mean ± SEM.
Analysis of leukocytes cell subpopulation in thymus
Analysis of thymus leukocyte subpopulations by flow cytometry. Thymuses were isolated from female C57BL/6 and B-hTSLP/hTSLPR mice plus (n=3, 9-week-old). Flow cytometry analysis of the thymus was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live cells were gated for the CD45+ population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of T cells, B cells, NK cells, dendritic cells, neutrophils, monocytes and macrophages in homozygous B-hTSLP/hTSLPR mice plus were similar to those in the C57BL/6 mice, demonstrating that TSLP, TSLPR and IL7R humanized does not change the overall development, differentiation or distribution of these cell types in thymus. Values are expressed as mean ± SEM.
Analysis of T cell subpopulation in thymus
Analysis of thymus T cell subpopulations by flow cytometry. Thymuses were isolated from female C57BL/6 and B-hTSLP/hTSLPR mice plus (n=3, 9-week-old). Flow cytometry analysis of the thymus was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live CD45+ cells were gated for CD3+ T cell population and used for further analysis as indicated here. B. Results of FACS analysis. The percent of CD4+ T cells, CD8+ T cells and Tregs in homozygous B-hTSLP/hTSLPR mice plus were similar to those in the C57BL/6 mice, demonstrating that TSLP, TSLPR and IL7R humanized does not change the overall development, differentiation or distribution of these T cell types in thymus. Values are expressed as mean ± SEM.
Functional analysis
Data from Huabo Biopharm Co. Ltd.
Mouse TARC was respectively induced with human TSLP and mouse TSLP in wild type C57BL/6 mice, homozygous B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus. Dendritic cells were respectively induced with FLT3L from bone marrow of the three strains and stimulated with human TSLP or mouse TSLP in vitro. Concentration of mouse TARC secreted from DCs was assayed by ELISA. Mouse TARC was successfully induced with human TSLP, but not mouse TSLP in homozygous B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus. The level of mTARC in B-hTSLP/hTSLPR mice plus was higher than that in B-hTSLP/hTSLPR mice plus. Meanwhile mouse TARC was successfully induced with mouse TSLP, but not human TSLP in wild-type C57BL/6 mice. Results indicated that TSLP and TSLPR are not cross-reactive between mouse and human. Human TSLP can activate the dendritic cells of B-hTSLP/hTSLPR mice and B-hTSLP/hTSLPR mice plus.
Growth curve
Growth curve of B-hTSLP/hTSLPR mice plus. Eight weeks old of mice were grouped (10 males and 10 females, respectively). Body weight was measured on the same day of every week and lasted for 12 weeks. The lowest and highest values of mouse body weight in the table were calculated from the mean ± SD. The growth curve conforms to the normal distribution and the probability of random error falling within ± SD is 68%.
Hematology analysis
Complete blood count (CBC) of B-hTSLP/hTSLPR mice plus. Values are expressed as mean ± SD.
Biochemistry analysis
Blood biochemical test of B-hTSLP/hTSLPR mice plus. Values are expressed as mean ± SD.
Gross anatomy of female B-hTSLP/hTSLPR mice plus
The organs of female B-hTSLP/hTSLPR mice plus (12-week-old, n=10).
Gross anatomy of male B-hTSLP/hTSLPR mice plus
The organs of male B-hTSLP/hTSLPR mice plus (12-week-old, n=10).
Organ weight
Average weight of the main organs of B-hTSLP/hTSLPR mice plus.
Histopathological analysis
In vivo efficacy of anti-human TSLP antibody with asthma mouse model
In vivo efficacy of anti-human TSLP antibody
Analysis of immune cells in BALF by FACS. Male B-hTSLP/hTSLPR mice plus were immunized with OVA to induce mouse asthma model (n=6). Anti-human TSLP antibody (Tezepelumab, synthesized in house) was given during the sensitization phase. BALF was collected at the end of the experiment to detect infiltrated inflammatory cells in lung tissue. The results showed that the proportion of eosinophils in OVA induced but untreated group (G2) was significantly higher than that in the non-induced group (G1), while the proportion of these cells in the treated group (G3) decreased significantly when compared with the G2 group. The number of CD45+ cells and eosinophils in the G3 group also tended to decline. Data indicated that anti-human TSLP antibodies could effectively reduce the number and proportion of eosinophils in B-hTSLP/hTSLPR mice plus induced with OVA.
In vivo efficacy of anti-human TSLP antibody
OVA specific IgE production in serum of mouse asthma model. Serum was collected at the study endpoint. IgE levels responded to OVA-specific antibody were analyzed. The results showed that the levels of IgE in mice treated with tezepelumab (in house) was lower than that in untreated mice.
In vivo efficacy of anti-human TSLP antibody
H&E staining of asthma-like model in B-hTSLP/hTSLPR mice plus
Lung tissues were collected at the study endpoint. H&E staining results showed that the lung tissues from B-hTSLP/hTSLPR mice plus exposed to PBS aerosols did not show any inflammation. OVA exposure resulted in a significant increase in peribronchial and perivascular inflammation in B-hTSLP/hTSLPR mice plus. A significantly reduction in eosinophils infiltration was observed in mice treated with tezepelumab (in house). Black arrow: inflammatory cells; black triangle: eosinophils.
Data to be released:
Concentration of total IgE in serum
Experimental schedule for Induction of AD-like skin lesions and in vivo efficacy of anti-human TSLP antibody
Experimental schedule for Induction of atopic dermatitis (AD)-like skin lesions and in vivo efficacy of anti-human TSLP antibody in B-hTSLP/hTSLPR mice plus.
OXA was applied to ear skin of mice on day 0, and then challenge to the same site of skin nine times from days 7 to 25.
Anti-human TSLP antibody tezepelumab (in house) was administered by intraperitoneal injection (n = 6). OXA: oxazolone.
In vivo efficacy of anti-human TSLP antibody in OXA induced AD-like mouse model
Efficacy of anti-human TSLP antibody in B-hTSLP/hTSLPR mice plus.
Mice in each group were treated with anti-hTSLP antibody tezepelumab (in house). (A) Statistical analysis of ear thickness in each group.
Epidermis of ear began to desquamate from day 18. So the ear thicknesses were decreased from day 18 as shown in figure.
(B) Body weight changes during the treatment. (C) Total IgE levels in serum. Serum was collected on day 26 and total IgE levels were measured by ELISA. (n = 6). Values are expressed as mean ± SEM.
H&E staining of ear skin in AD-like mouse model of B-hTSLP/hTSLPR mice plus
Effects of anti-human TSLP antibody on ear skin of the AD mouse model.
(A) Hematoxylin and eosin (H&E) staining. (B) Thickness of ear epidermal skin. (C) Score of eosinophils infiltrated in ear epidermal skin.
(D) Total score of ear epidermal skin.
Ear thickness and infiltration scores of eosinophils in ear skin of the groups treated with Tezepelumab (in house) were decreased significantly compared to that in the isotype control,
demonstrating that the B-hTSLP/hTSLPR mice plus provide a powerful preclinical model for in vivo evaluation of anti-human TSLP antibodies.
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. AD: Atopic dermatitis.
