by (Keying Zhu); resources, H.L., K.B. microglia in the mouse brain [7]. In addition, treatment of of liver tissue macrophages can be reduced using both mice, with control mice receiving a normal diet. Mice were terminated 18 or 21 days following PLX3397 treatment. No obvious side-effects were observed with PLX3397, except for whitening of fur color as previously reported [21]. Flow cytometric analyses of brain tissues were performed at each time point. Our results exhibited that CD11b+CD45lowLy6C?Ly6G? microglia can be effectively depleted 18 days (79.82% 4.45%) and 21 days (95.81% 1.91%) following PLX3397 treatment at a concentration of 290 mg/kg (Physique 1ACC, **** 0.0001). Open in a separate windows Physique 1 Microglia are effectively CBL-0137 depleted 21 days following PLX3397 treatment. (A) Representative flow cytometry plots of CD11b+CD45lowLy6C?Ly6G? microglia of the hemi-brains in mice following consecutive PLX3397 diet Igf1 (18 and 21 days) at a dose of 290 mg/kg. Control mice were treated with normal diet. (B) Total CD11b+CD45lowLy6C?Ly6G? microglial counts ( SEM) of the hemi-brains during microglial depletion periods (control, black bars; day 18, blue bars; day 21, red bars). (C) Percentages of CD11b+CD45lowLy6C?Ly6G? microglia (/control, SEM) during microglial depletion periods (control, black bars; day 18, blue bars; day 21, red bars, = 11, 6, 4, respectively). Statistical significance is usually indicated as **** 0.0001. 2.2. Splenic Red Pulp Macrophages Are Significantly Decreased Following Both Conditional Genetic and Pharmacological Microglial Depletion We next resolved whether splenic red pulp macrophages were influenced after conditional genetic and pharmacological microglial depletion. 0.05, ** 0.01). Comparable findings with reduced numbers and percentages of F4/80hiLy6C? splenic red pulp macrophages were also noted using 290 mg/kg PLX3397 diet pharmacological-induced microglial depletion. Our results discerned that this numbers and percentages of F4/80hiLy6C? red pulp macrophages in the spleen were significantly fewer 21 days following PLX3397 treatment, the most significant time point of pharmacological microglial depletion, than control group (Physique 2C,D ** 0.01). Open in a separate window Physique 2 Splenic red pulp macrophages are reduced following both conditional genetic and pharmacological microglial depletion. (A) Representative flow cytometry plots of splenic red pulp macrophages in = 6, 4, 4, 6, 6, respectively). (C) Representative flow cytometry plots of splenic red pulp macrophages CBL-0137 in mice following 290 mg/kg PLX3397 diet (21 days). Control mice were treated with normal diet. (D) The numbers and percentages of splenic red pulp macrophages ( SEM) during pharmacological microglial depletion periods (control, black bars; day 21, red bars, = 10, 4, respectively). A total of 400,000 cells from the spleen were run for flow cytometry in this experiment. Statistical significance is usually indicated as * 0.05 and ** 0.01 (ns: not significant). 2.3. Splenic Ly6Chi Monocytes Are Significantly Increased Following Conditional Genetic Microglial Depletion, but Not Pharmacological Depletion We next resolved whether splenic Ly6Chi monocytes could also be influenced following both genetic and pharmacological microglial depletion. Unlike reduced numbers of splenic red pulp macrophages, we exhibited that during conditional genetic microglial depletion periods the numbers and percentages of splenic Ly6Chi monocytes were significantly increased 7 days following tamoxifen CBL-0137 treatment (Physique 3A,B **** 0.0001; ** 0.01). The numbers and percentages of splenic Ly6Chi monocytes were not significantly different between = 6, 4, 4, 6, 6, respectively). (C) Representative flow cytometry plots of splenic Ly6Chi monocytes in mice following PLX3397 diet (21 days) at a dose of 290 mg/kg. Control mice were treated with normal diet. (D) The numbers and percentages of splenic Ly6Chi monocytes ( SEM) during pharmacological microglial depletion periods (control, black bars; day 21, red bars, = 11, 4, respectively). Statistical significance is usually indicated as ** 0.01 and **** 0.0001 (ns: not significant). 2.4. Splenic CD4+ T Cells and NK Cells Are Affected Following Conditional Genetic and Pharmacological Microglial Depletion Our results exhibited that during both conditional genetic and pharmacological microglial depletion periods the numbers of splenic CD4+ T cells were significantly decreased after 7 days (Physique 4A,B *** 0.001) and 21 days (Physique 4D,E * 0.05), respectively. Furthermore, the numbers and percentages of splenic NK cells were significantly reduced following both conditional genetic and pharmacological microglial depletions (Physique 4C,F **** 0.0001, * 0.05). Open in a separate window Physique 4 Splenic CD4+ T cells and NK cells are affected following conditional genetic and pharmacological microglial depletion. (A) Representative flow cytometry plots of splenic CD4+ T cells and.
