TNFAIP3 deletion is induced using conditional knock-out mice for myeloid lineage. deletion in myeloid cells induces a reduction in body weight, a decrease in the number of M-MDC and of common monocyte and granulocyte precursor cells (CMGPs). We also reported that the lack of TNFAIP3 in myeloid cells induces an increase in microglial cell density. The results suggest that TNFAIP3 in myeloid cells critically controls the development of M-MDC in lymphoid organ and of microglia in the CNS. = 7) showed a lower body weight compared to their WT littermates (= 7) (Figure 1, MannCWhitney test = 0.014). No others macroscopic abnormalities were observed in TNFAIP3cx3cr1-KO mice in comparison to their WT littermates. Open in a separate window Figure 1 Body weight analysis of 3 month-old WT and TNFAIP3cx3cr1-KO mice. The analysis of body weight reports that at 3 months of age, TNFAIP3cx3cr1-KO mice are smaller compared to their WT littermates (= 7 for each group) (MannCWhitney test, = 0.014 * 0.05). To characterize the immunological phenotype of TNFAIP3cx3cr1-KO mice due to the absence of TNFAIP3 gene, the flow-cytometry analysis on samples obtained from spleen (Figure 2 and Figure 3 and Table S1) and lymph nodes (Figure 2 and Figure 4 and Table S1) of adult Mouse monoclonal antibody to ACE. This gene encodes an enzyme involved in catalyzing the conversion of angiotensin I into aphysiologically active peptide angiotensin II. Angiotensin II is a potent vasopressor andaldosterone-stimulating peptide that controls blood pressure and fluid-electrolyte balance. Thisenzyme plays a key role in the renin-angiotensin system. Many studies have associated thepresence or absence of a 287 bp Alu repeat element in this gene with the levels of circulatingenzyme or cardiovascular pathophysiologies. Two most abundant alternatively spliced variantsof this gene encode two isozymes-the somatic form and the testicular form that are equallyactive. Multiple additional alternatively spliced variants have been identified but their full lengthnature has not been determined.200471 ACE(N-terminus) Mouse mAbTel+ 3 month old WT (= 8) and TNFAIP3cx3cr1-KO (= 4) mice was performed. Notably, the analysis on the spleens highlighted that the percentage number of macrophages (Figure 3A, MannCWhitney test = 0.004), monocytes (Figure 3B, MannCWhitney test = 0.016), DCs (Figure 3C, MannCWhitney test = 0.048) and B cells (Figure 3F, MannCWhitney test = 0.048) was reduced in TNFAIP3cx3cr1-KO mice compared to their WT littermates. No differences were reported in the other cellular populations analyzed, such as natural killer (NK) (Figure 3D, MannCWhitney test = 0.214), NK T (Figure 3E, MannCWhitney test = 0.153), CD3+ T lymphocytes (Number 3G, MannCWhitney test = 0.367), CD4+ T helper (Number 3H, MannCWhitney test VXc-?486 = 0.683) and CD8+ T cytotoxic (Number 3I, MannCWhitney test = 0.109) cells. These results indicate the deletion of TNFAIP3 in myeloid cells prospects to an modified immunological phenotype not only in myeloid linage as expected but also in lymphocytes, such as B cells. Open in a separate window Open in a separate window Number 2 Flow-cytometry gating strategy utilized for spleen and lymph nodes. (A) Gating strategy for lymphocytes, NK and B cells; (1) VXc-?486 FS versus SS gating was used to discriminate cells based on their size; (2) living cells were then recognized by propidium iodide negativity; (3) CD3 was used like a T-cell marker; (4) CD3+ cells were distinguished based on the presence of CD4 and CD8; (5) NK and NK T were distinguished based on the presence of CD49b; (6) CD45R was used like a B-cell marker. (B) Gating strategy for macrophages, monocytes and DCs; (1) FS versus SS gating was used to discriminate cells based on their size; (2) living cells were then recognized by propidium iodide negativity; (3C4) F4/80 gating on CD11b+ cells was used to select macrophages; (5) Ly6-C and Ly6-G gating on CD11b+ cells was used to select monocytes; (6) DCs were distinguished based on the presence of CD11c and CD86. (NK, natural killer; FS, ahead scatter; SS part scatter; DCs, dendritic cells). Open in a separate window Number 3 Flow-cytometry analysis performed on spleen from 3 month-old WT and TNFAIP3cx3cr1-KO mice. The analysis reveals the percentage quantity of CD11b+F4/80+ macrophages (A, = 0.004), CD11b+Ly-6C+Ly-6G+ monocytes VXc-?486 (B, = 0.016), CD11c+CD86+ DCs (C, = 0.048) and B200+ B (F, = 0.048) cells is reduced in TNFAIP3cx3cr1-KO mice compared to their WT littermates. No variations are reported in CD49+NK (D, = 0.214) and CD3+ CD49+NK T cells (E, = 0.153) CD3+ T (G, = 0.367), CD3+CD4+ T helper (H, = 0.683) and CD3+CD8+ cytotoxic T cells (I, = 0.109) (WT = 8; TNFAIP3cx3cr1-KO = 4) (MannCWhitney test, ** 0.01; * 0.05). (DCs, dendritic cells; NK, natural killer). Open in a separate window Number 4 Flow-cytometry analysis performed on lymph nodes from 3 month-old WT and TNFAIP3cx3cr1-KO mice. The analysis reveals the percentage quantity of CD11b+F4/80+ macrophages (A, = 0.006), CD49+ NK (D, = 0.009) and CD3+CD49+ NK T (E, = 0.009) cells is definitely reduced in TNFAIP3cx3cr1-KO mice compared to their WT littermates. No variations are reported in CD11b+Ly-6C+Ly-6G+ monocytes (B, = 0.914), CD3+ T (G, = 0.114) and CD3+CD4+ T.
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