B-NDG hIL6 mice
Il6 (interleukin 6）
IL6 (interleukin 6）this gene encodes a cytokine that functions in inflammation and the maturation of B cells. The encoded protein has been shown to be an endogenous pyrogen capable of inducing fever in people with autoimmune diseases or infections. Humanized mice are a powerful tool for the study of human hematopoiesis and immune function in vivo. However, the existing models cannot support robust adaptive immune responses, especially the generation of class-switched, antigen-specific antibody responses . Interleukin 6 (IL-6) was initially identiﬁed as a B-cell differentiation factor both in vivo and in vitro. It is capable of inducing the ﬁnal at uration of B cells into immunoglobulin-secreting plasma cells .In fact, IL-6 has been shown to stimulate the secretion of antibodies to such a degree that serum IgG1 levels can rise 120- to 400-fold .Because murine and human IL-6 show 65% sequence homology at the DNA level and 42% homology at the protein level ,16 and murine IL-6 is not active in human cells,we reasoned that physiological expression of human IL-6 in the mouse may result in improved B-cell differentiation and human antibody production.
Biocytogen developed the B-NDG hIL6 mice, and the targeting strategy was that to replace the sequence encompassing the ORF of mouse Il6 with its human counterpart. This mice combines a B-NDG mouse background and expresses human IL6 cytokine, not only improves thymopoiesis and peripheral T-cell engraftment,but also signiﬁcantly increases the level of total IgG and antigen-speciﬁc IgG, will become a suitable animal model to investigate human immune system.
Generation of gene editing mouse model and expression analysis
Gene targeting strategy for B-NDG hIL6 mice. The Il6 sequence encompassing the ORF of mouse were replaced by human counterpart (a 4.8 kb human IL6 sequence extending from ATG in exon 1 to exon 5 with 16 nucleotides of 3’ downstream sequence), maintain the promoter and other regulatory elements of mouse origin in B-NDG hIL6 mice.
Protein expression analysis (heterozygous mice)
Species specific IL6 expression analysis in B-NDG hIL6 mice by ELISA. Serum from B-NDG and heterozygous B-NDG hIL6 (H/+) mice were analyzed ELISA with anti-IL6 antibodies. Mouse IL6 was detectable in B-NDG and heterozygous B-NDG hIL6 mice. Human IL6 were exclusively detectable in heterozygous B-NDG hIL6 but not B-NDG mice.
B-NDG hIL6 mice are well suited for human immune system engraftment
Human CD34+ cells(0.15M) were intravenous implanted into homozygote B-NDG hIL6 and B-NDG mice (female, 6 week-old, n=15). All mice were treated with 1.6gry-irradiation. Representative flow cytometric analysis of PBMCs from mice after engraftment with human CD34+ cells. B-NDG hIL6 show a higher percentage of human myeloid cells and monocyte compared with B-NDG.
Human CD34+ cells(0.15M) were intravenous implanted into homozygote B-NDG hIL6 and B-NDG mice (female, 6 week-old, n=14). All mice were treated with 1.6gry-irradiation. Representative flow cytometric analysis of PBMCs from mice after engraftment with human CD34+ cells. B-NDG hIL6 show a higher percentage of human leukocytes, myeloid cells and monocyte compared with B-NDG.