• IL23 is a member of IL-12-type cytokine family controlling Th17 cells development. Excessive IL-23 signaling has been implicated in the pathogenesis of autoimmune diseases. IL23 is a heterodimer consisting of the unique subunits p19 and p40. IL23 receptor complex consist of two subunits: IL12Rβ1 and IL23R. IL23R is the unique IL23 receptor subunit and IL-12Rβ1 is also the IL12 receptor complex.
• The IL23-IL23R complex signals through JAK kinase and STAT transcription factors, which activates Jak2 and Tyk2 kinases, phosphorylates the receptor, leading to phosphorylation of STAT3, thus activates several pathways including IL17A, IL17F and IL22.
• Gene editing strategy: The extracellular region sequences of mouse Il12rb1 were replaced by human IL12RB1 counterpart gene sequences in B-hIL23R/hIL12RB1 plus mice. Chimeric CDS, composed of human IL23R extracellular region plus mouse Il23r transmembrane and cytoplasmic region, was inserted into the mouse Il23r gene exon 3 in B-hIL23R/hIL12RB1 plus mice.
• mRNA expression analysis: Mouse Il12rb1 mRNA was only detectable in wild-type mice. Human IL12RB1 mRNA was exclusively detectable in B-hIL23R/hIL12RB1 plus mice (H/+, H/H). Mouse Il23r mRNA was detectable in wild-type mice. Human IL23R mRNA was exclusively detectable in homozygous B-hIL23R/hIL12RB1 plus mice (H/H, H/H).
• Functional analysis: Human IL23 can induced mouse IL17A production in wild-type C57BL/6JNifdc mice and homozygous B-hIL23R/hIL12RB1 plus mice. The positive drug can inhibit the expression of mouse IL17A in homozygous B-hIL23R/hIL12RB1 plus mice with hIL23 stimulated.
• Leukocytes cell subpopulation analysis: Humanization of IL23R and IL12RB1 does not change the overall frequency or distribution of immune cell types in spleen, blood and lymph nodes. Application: This product is used for pharmacodynamics and safety evaluation of  tumors and autoimmune diseases.

Gene targeting strategy for B-hIL23R/hIL12RB1 plus mice. 

• The extracellular region sequences of mouse Il12rb1 were replaced by human IL12RB1 counterpart gene sequences in B-hIL23R/hIL12RB1 plus mice. 
• Chimeric CDS, composed of human IL23R extracellular region plus mouse Il23r transmembrane and cytoplasmic region, was inserted into the mouse Il23r gene exon 3 in B-hIL23R/hIL12RB1 plus mice.

Strain specific analysis of IL12RB1 mRNA expression in wild-type C57BL/6JNifdc mice and B-hIL23R/hIL12RB1 plus mice by RT-PCR. Spleen separated CD4+ T cells RNA were isolated from wild-type C57BL/6JNifdc mice (+/+) and B-hIL23R/hIL12RB1 plus mice (H/+, H/H), and then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human IL12RB1 primers. Mouse Il12rb1 mRNA was only detectable in wild-type mice. Human IL12RB1 mRNA was exclusively detectable in B-hIL23R/hIL12RB1 plus mice.

Strain specific analysis of IL23R mRNA expression in wild-type C57BL/6JNifdc mice and B-hIL23R/hIL12RB1 plus mice by RT-PCR. Splenocytes RNA were isolated from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hIL23R/hIL12RB1 plus mice (H/H, H/H), and then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human IL23R primers. Mouse Il23r mRNA was detectable in wild-type mice. Human IL23R mRNA was exclusively detectable in homozygous B-hIL23R/hIL12RB1 plus mice.

Function analysis of IL23R in wild-type C57BL/6JNifdc mice and homozygous B-hIL23R/hIL12RB1 plus mice by ELISA. Splenocytes were collected from wild-type C57BL/6JNifdc (+/+) mice and homozygous B-hIL23R/hIL12RB1 plus mice (H/H), then CD4+ T cells were isolated and cultured on anti-mCD3ε antibody and anti-mCD28 antibody pre-coated plates, and stimulated with indicated concentrations of mouse IL23 recombinant protein (10 ng/mL) or human IL23 recombinant protein (10 ng/mL), and different concentrations of positive drug (0 nM, 0.2 nM, 1 nM) at 37℃ for 48 h. Cell supernatants were collected for ELISA analysis of mouse IL17A (BioLegend, 432504). Mouse IL23 can induced mouse IL17A production in wild-type C57BL/6JNifdc mice and homozygous B-hIL23R/hIL12RB1 plus mice. Human IL23 can induced mouse IL17A production in wild-type C57BL/6JNifdc mice and homozygous B-hIL23R/hIL12RB1 plus mice. The positive drug can inhibit the expression of mouse IL17A in homozygous B-hIL23R/hIL12RB1 plus mice with hIL23 stimulated. The positive drug is provided by the client.

Frequency of leukocyte subpopulations in spleen by flow cytometry. Splenocytes were isolated from wild-type C57BL/6JNifdc mice (male, n=3, 6-week-old) and homozygous B-hIL23R/hIL12RB1 plus mice (male, n=3, 6-week-old). 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, granulocytes, monocytes, macrophages, CD4+ T cells, CD8+ T cells and Tregs in B-hIL23R/hIL12RB1 plus mice were similar to those in C57BL/6JNifdc mice, demonstrating that humanization of IL23R and IL12RB1 does not change the frequency or distribution of these cell types in spleen. The frequency of leukocyte subpopulations in lymph nodes and blood of B-hIL23R/hIL12RB1 plus mice were also comparable to wild-type C57BL/6JNifdc mice (Data not shown). Values are expressed as mean ± SEM.