Figure 7.

Antibiotics administration reduces CTX-driven myeloid cell expansion and the suppressive activity of monocytes. (A) CTX activates TLR4 signaling pathway in monocytes. Naïve BALB/c mice were treated with CTX. On day 7, CD11b⁺Ly6C^hi monocytes were flow-sorted from the splenocytes recovered from CTX-treated mice. Monocytes from the splenocytes from naïve BALB/c mice were used as control. Total RNA was extracted from sorted monocytes and subjected to qRT-PCR to evaluate the mRNA levels of the indicated genes. The target gene transcripts are normalized to β-actin. (B) Antibiotics administration inhibits CTX-induced myeloid cell expansion. Normal drinking water (H₂O) or antibiotics-containing water (ABX) was provided to mice 7 days before CTX treatment, and maintained for the duration of the experiment. 7 days after CTX treatment, spleen cells were processed and stained for CD11b and Ly6C. Representative dot plots are shown, and the numbers indicate the frequencies of gated populations. The frequencies of each myeloid subset are summarized in bar graphs with at least three mice each group. (C) The bar graphs summarize the numbers of monocytes and neutrophils in mice receiving CTX or CTX+ABX treatment. (D) ABX administration reduces the suppressive activity of monocytes from CTX-treated mice. Violet dye-labeled responder T cells were mixed with indicated numbers of sorted myeloid cells derived from mice receiving either CTX only (-ABX) or CTX+ABX (+ABX), and stimulated with aCD3/aCD28 mAbs. Proliferation of T cells was measured by violet dye dilution. Numbers in histogram indicate the percent of undivided responder T cells. *p < 0.05, **p < 0.01, ***p < 0.001.