• RAGE is expressed in more than 300 cells in addition to lung tissue, and studies have shown that tumors can increase the expression of this gene in a variety of cells.
• Exons 1-11 including 3’UTR of the mouse RAGE gene that encodes the full-length protein was replaced by human RAGE exons 1-11 including 3’UTR in B-hRAGE mice plus.
• Mouse RAGE protein was only detectable in wild-type C57BL/6 mice. Human RAGE protein was detectable in homozygous B-hRAGE mice plus but not in wild-type mice.
• Mouse RAGE mRNA was only detectable in wild-type C57BL/6 mice. Human RAGE mRNA was detectable in homozygous B-hRAGE mice plus but not in wild-type mice.

Gene targeting strategy for B-hRAGE mice plus. Exons 1-11 including 3’UTR of the mouse RAGE gene that encodes the full-length protein was replaced by human RAGE exons 1-11 including 3’UTR in B-hRAGE mice plus.

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

Strain specific analysis of RAGE expression in wild-type C57BL/6N mice and heterozygous humanized B-hRAGE mice plus by RT-qPCR. Blood, and blood RNA were isolated from wild-type C57BL/6N mice (+/+) (n=3, 9-week-old) and B-hRAGE mice plus (n=5, 9-week-old), then cDNA libraries were synthesized by reverse transcription, followed by real-time quantitative PCR with RAGE primers. The mRNA levels of RAGE was in detectable in B-hRAGE mice plus with Applied Biosystems SYBR, but not wild-type C57BL/6N mice. The mRNA levels of RAGE in lung from B-hRAGE mice plus was higher than that in blood from B-hRAGE mice plus.

Western blot analysis of RAGE protein expression in homozygous B-hRAGE mice plus. Lung tissue lysates were collected from wild-type C57BL/6N mice (+/+), homozygous B-hRAGE mice plus (H/H) and B-Rage KO mice(RM), and then analyzed by western blot with species-specific anti-RAGE antibody. 40 μg total proteins were loaded for western blotting analysis. RAGE was detected in lung.

Strain specific RAGE expression analysis in wild-type C57BL/6N mice and heterozygous humanized B-hRAGE mice plus by ELISA. Serum was collected from wild-type C57BL/6N mice (+/+) (male, n=3, 7-week-old) and heterozygous B-hRAGE mice plus (H/+) (male, n=3, 15-week-old). Expression level of mouse and human RAGE were analyzed by ELISA (Human RAGE Quantikine ELISA Kit: R&D, DRG00). Human RAGE was exclusively detectable in heterozygous B-hRAGE mice plus(n=3). Values are expressed as mean ± SEM.

Strain specific RAGE expression analysis in wild-type C57BL/6N mice and heterozygous humanized B-hRAGE mice plus by ELISA. Lung homogenates were collected from wild-type C57BL/6N mice (+/+) (male, n=3, 7-week-old) and heterozygous B-hRAGE mice plus (H/+) (male, n=3, 15-week-old). Expression level of mouse and human RAGE were analyzed by ELISA (Human RAGE Quantikine ELISA Kit: R&D, DRG00). Human RAGE was exclusively detectable in heterozygous B-hRAGE mice plus(n=3). Values are expressed as mean ± SEM.

The inhibitory efficiency of the RAGE targeted nucleic acid drugs in homozygous B-hRAGE mice plus. B-hRAGE mice plus were randomly divided into 3 groups (5-6 weeks old, female). The human RAGE-targeted nucleic acid drug (provided by a client) and PBS were administered to the mice individually. The nucleic acid drug was administered in the form of PBS aqueous solution. The mice were sacrificed on day 7. (A) The schematic diagram of experimental processing. (B) The changes in RAGE mRNA expression levels in lung tissue on day 7 after administration, compared to the levels before administration. Values are expressed as mean ± SEM.