Basic Information
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Targeting strategy
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Gene targeting strategy for B-hIL36R/hIL1RACP mice.
The chimeric IL1RL2 CDS was inserted after the 5’UTR of mouse Il1rl2 gene in B-hIL36R/hIL1RACP mice. The insertion disrupts the endogenous murine Il1rl2 gene, resulting in the absence of mouse transcripts.
The chimeric IL1RAP CDS was inserted after the 5’UTR of mouse Il1rap gene in B-hIL36R/hIL1RACP mice. The insertion disrupts the endogenous murine Il1rap gene, resulting in the absence of mouse transcripts.
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mRNA expression analysis
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Strain specific analysis of IL36R gene expression in wild type (WT) mice, B-hIL36R mice and B-hIL36R/hIL1RACP mice by RT-PCR.
Human IL36R mRNA was detectable only in lung of homozygous B-hIL36R mice (H/H) and homozygous B-hIL36R/hIL1RACP mice (H/H;H/H) but not in WT mice (+/+). The positive band was confirmed to be correct by sequencing.
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Protein expression analysis
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Strain specific IL1RACP expression analysis in homozygous B-hIL36R/hIL1RACP mice by flow cytometry. Monocytes, macrophages and neutrophils were collected from wild type (WT) mice (+/+) and homozygous B-hIL36R/hIL1RACP mice (H/H;H/H), and analyzed by flow cytometry with species-specific anti-hIL1RACP antibody. Human IL1RACP was exclusively detectable in homozygous B-hIL36R/hIL1RACP mice (H/H;H/H) but not in WT mice (+/+).
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Blood routine test in B-hIL36R/hIL1RACP mice
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Complete blood count (CBC). Blood from female C57BL/6 and B-hIL36R/hIL1RACP mice (n=8, 8 week-old) was collected and analyzed for CBC. There was no differences among any measurement between C57BL/6 and B-hIL36R/hIL1RACP mice, indicating that introduction of IL36R and IL1RACP in place of its mouse counterpart do not change blood cell composition and morphology. Values are expressed as mean ± SEM.
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Blood biochemistry of B-hIL36R/hIL1RACP mice
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Blood biochemistry tests of B-hIL36R/hIL1RACP mice. Serum from the C57BL/6 and B-hIL36R/hIL1RACP mice (n=8, 8 week-old) was collected and analyzed for levels of ALT and AST. There was no differences on either measurement between C57BL/6 and B-hIL36R/hIL1RACP mice, indicating that introduction of IL36R and IL1RACP in place of its mouse counterpart do not change ALT and AST levels or health of liver. Values are expressed as mean ± SEM.
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IL36g-induced skin inflammation in B-hIL36R/hIL1RACP mice
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Homozygous B-hIL36R/hIL1RACP mice were induced ear skin inflammation with injection of human IL36g cytokine (2ug/25uL per dose) intradermal in ear skin, and pre-treated with IL-36 pathway blocker Spesolimab i.v. (10 mg/kg), and measuring ear skin thickness. The results showed that human IL-36g cytokine induced the ear skin inflammation significantly in B-hIL36R/hIL1RACP mice as the ear skin thickening, and IL-36 pathway blocker Spesolimab could relieve skin inflammation. Values are expressed as mean ± SEM. The data was obtained from a partner.
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Summary
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- mRNA expression analysis:
Human IL36R mRNA was detectable only in lung of homozygous B-hIL36R mice (H/H) and homozygous B-hIL36R/hIL1RACP mice (H/H;H/H) but not in WT mice (+/+). The positive band was confirmed to be correct by sequencing.
- Protein expression analysis:
Human IL1RACP was exclusively detectable on monocytes, macrophages and neutrophils of homozygous B-hIL36R/hIL1RACP mice (H/H;H/H) but not in WT mice (+/+).
- Blood routine test and Blood biochemical test
IL36R and IL1RACP humanized do not change the blood cell composition and morphology, ALT and AST levels or health of liver.
- In vivo efficacious：
Human IL-36 cytokine induced the ear skin inflammation significantly in B-hIL36R/hIL1RACP mice as the ear skin thickening. And the IL-1 and IL-36 pathway blockers could relieve skin inflammation.
