• CD11b is an alpha chain of the integrin receptor CD11b/CD18 (also known as aM2, Mac-1, and CR3), encoded by the ITGAM gene and belonging to the integrin β2 family. CD11b is mainly expressed in monocytes, macrophages, dendritic cells and granulocytes. CD11b, whose biological functions include both pro-inflammatory and anti-inflammatory effects, binds to a variety of ligands, including members of the intercellular adhesion molecule family, complement protein C3b and fibrinogen, and mediates leukocyte adhesion, migration, and tissue recruitment. CD11b is involved in many pro-inflammatory biological processes primarily related to infection control. Other studies have shown that CD11b knockout can promote tumor growth, increase the expression of immunosuppressive cytokines, and promote the differentiation of macrophages into immunosuppressive macrophages. CD11b/CD18 small molecule agonist activation can regulate the differentiation of macrophages into pro-inflammatory macrophages, and promote the recruitment of CD8+T cells into tumor tissues. Finally, the purpose of inhibiting tumor growth is achieved.
• mRNA expression analysis: Mouse Cd11b was only detectable in wild-type C57BL/6JNifdc mice. Human CD11B was only detectable in homozygous B-hCD11B mice.
• Protein expression analysis: CD11B was detectable in wild-type C57BL/6JNifdc mice and homozygous B-hCD11B mice, as anti-mouse/human CD11B antibody was cross-reactive between mouse and human. Human CD11B was only detectable in homozygous B-hCD11B mice. Mouse CD11B was detectable in NK cells, DCs, neutrophils, monocytes and macrophages of wild-type mice. Human CD11B was detectable in neutrophils, but weakly detectable in NK cells, DCs, monocytes and macrophages.

Gene targeting strategy for B-hCD11B mice.
The exons 2-29 of mouse Itgam gene that encode extracellular domain are replaced by human counterparts in B-hCD11B mice. The genomic region of mouse Itgam gene that encodes signal peptide, transmembrane domain and cytoplasmic portion is retained. The promoter, 5’UTR and 3’UTR region of the mouse gene are also retained. The chimeric CD11B expression is driven by endogenous mouse Itgam promoter, while mouse Itgam gene transcription and translation will be disrupted.

Strain specific analysis of CD11B mRNA expression in wild-type C57BL/6JNifdc mice and B-hCD11B mice by RT-PCR. Spleen RNA were isolated from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hCD11B mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human CD11B primers. Mouse Cd11b mRNA was detectable only in wild-type mice. Human CD11B mRNA was detectable only in homozygous B-hCD11B mice but not in wild-type mice.

Strain specific CD11B expression analysis in wild-type mice and homozygous B-hCD11B mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6JNifdc mice (+/+) and B-hCD11B mice (H/H) (male, 9-week-old, n=1), and analyzed by flow cytometry with anti-mouse or human CD11B antibody (Biolegend, 101243) and anti-human CD11B antibody (Biolegend, 301306). Mouse CD11B was detectable in NK cells, but not in B cells and T cells of wild-type mice. Human CD11B was detectable in NK cells, but not in B cells and T cells of homozygous B-hCD11B mice.

Strain specific CD11B expression analysis in wild-type mice and homozygous B-hCD11B mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6JNifdc mice (+/+) and B-hCD11B mice (H/H) (male, 9-week-old, n=1), and analyzed by flow cytometry with anti-mouse or human CD11B antibody (Biolegend, 101243) and anti-human CD11B antibody (Biolegend, 301306). Mouse CD11B was detectable in DCs, neutrophils, monocytes and macrophages of wild-type mice. Human CD11B was detectable in neutrophils, monocytes and macrophages, but weakly detectable in DCs of homozygous B-hCD11B mice.

Strain specific CD11B expression analysis in wild-type mice and homozygous B-hCD11B mice by flow cytometry. Blood cells were collected from wild-type C57BL/6JNifdc mice (+/+) and B-hCD11B mice (H/H) (male, 9-week-old, n=1), and analyzed by flow cytometry with anti-mouse or human CD11B antibody (Biolegend, 101243) and anti-human CD11B antibody (Biolegend, 301306). Mouse CD11B was detectable in NK cells, but not in B cells and T cells of wild-type mice. Human CD11B was weakly detectable in NK cells, but not in B cells and T cells of homozygous B-hCD11B mice.

Strain specific CD11B expression analysis in wild-type mice and homozygous B-hCD11B mice by flow cytometry. Blood cells were collected from wild-type C57BL/6JNifdc mice (+/+) and B-hCD11B mice (H/H) (male, 9-week-old, n=1), and analyzed by flow cytometry with anti-mouse or human CD11B antibody (Biolegend, 101243) and anti-human CD11B antibody (Biolegend, 301306). Mouse CD11B was detectable in DCs, neutrophils, monocytes and macrophages of wild-type mice. Human CD11B was detectable in neutrophils, but weakly detectable in DCs, monocytes and macrophages of homozygous B-hCD11B mice.