Figure 3. Effects of TBI, chemotherapy, commensal bacteria, and uric acid on IL-1β/TNF production. (A and B) RNA from the intestines and skin of mice receiving TBI (9 Gy) or chemotherapy (BU/Cyclophosphamide) was isolated 24 h after conditioning, and the expression of IL-1β and TNF was determined by quantitative PCR (qPCR). Neomycin (neo) or Ciprofloxacin (CPFL) were given starting 7 d before TBI, as indicated. Each data point represents an individual animal (n = 3) and the experiment was repeated 3 times. (C, left) Intestinal tract cells were isolated on d7 after allo-HCT, and the amount of intracellular IL-1β production was determined in the indicated cell populations after stimulation with PMA/ionomycin for 3 h. Shown are pooled results of six mice per group. (C, right) C57BL/6 hosts were transplanted (TBI, BALB/c BM+ T cells) and sacrificed 3 or 7 d after transplantation or left untreated (steady state). LPLs were isolated and directly analyzed for expression of CD11b, CD11c, donor/recipient markers, and intracellular expression of pro–IL-1β by flow cytometry. Top graphs show myeloid subgating of live LPLs, bottom graphs show donor H2k expression versus pro–IL-1β of myeloid subpopulations from top row. Data from one of two independent experiments with similar results and multiple time point analyses are shown. (D) RNA from the intestines of mice receiving TBI was isolated after 24 h and the expression of IL-1β was determined by quantitative PCR (qPCR). Each data point represents an individual animal (n = 4). The experiment was performed 3 times with similar results. (E) C57BL/6 mice received anti–IL-1β antibody or vehicle 24h before TBI or control treatment. RNA from the small intestine was isolated 40 h after TBI, and the expression of IL-1β was determined by quantitative PCR (qPCR). Each data point represents an individual animal (n = 4). The experiment was performed three times, with similar results. (F) BALB/c mice underwent different conditioning regimes: TBI alone (day 0; n = 3) or followed by allo-HCT with C57/BL6 BM plus T cells (day 10; n = 4); BU/CY chemotherapy or FLU/CY chemotherapy alone (day 0, n = 5) or followed by allo-HCT with C57/BL6 BM plus T cells (day 3, n = 6) as indicated. Untreated mice (n = 9) served as control. At the indicated time points, peritoneal fluid was isolated and UA levels were determined. Each data point represents an individual animal, and the experiment was performed twice with similar results. (G) BALB/c mice underwent TBI followed by allo-HCT with C57/BL6 BM alone (n = 10) or C57/BL6 BM plus T cells. Mice were treated with PBS (n = 16) or uricase starting from day −1 forward or from day +5 forward (n = 10 per group), as indicated. The experiment was performed twice and the resulting survival data were pooled. (H) On day 3 after allo-HCT, serum was isolated from BALB/c mice that received treatment with PBS or uricase from day 0 forward. Serum IL-1β levels were determined by ELISA. ***, P = 0.0001; **, P < 0.01; *, P < 0.05.
Figure 3.

Effects of TBI, chemotherapy, commensal bacteria, and uric acid on IL-1β/TNF production. (A and B) RNA from the intestines and skin of mice receiving TBI (9 Gy) or chemotherapy (BU/Cyclophosphamide) was isolated 24 h after conditioning, and the expression of IL-1β and TNF was determined by quantitative PCR (qPCR). Neomycin (neo) or Ciprofloxacin (CPFL) were given starting 7 d before TBI, as indicated. Each data point represents an individual animal (n = 3) and the experiment was repeated 3 times. (C, left) Intestinal tract cells were isolated on d7 after allo-HCT, and the amount of intracellular IL-1β production was determined in the indicated cell populations after stimulation with PMA/ionomycin for 3 h. Shown are pooled results of six mice per group. (C, right) C57BL/6 hosts were transplanted (TBI, BALB/c BM+ T cells) and sacrificed 3 or 7 d after transplantation or left untreated (steady state). LPLs were isolated and directly analyzed for expression of CD11b, CD11c, donor/recipient markers, and intracellular expression of pro–IL-1β by flow cytometry. Top graphs show myeloid subgating of live LPLs, bottom graphs show donor H2k expression versus pro–IL-1β of myeloid subpopulations from top row. Data from one of two independent experiments with similar results and multiple time point analyses are shown. (D) RNA from the intestines of mice receiving TBI was isolated after 24 h and the expression of IL-1β was determined by quantitative PCR (qPCR). Each data point represents an individual animal (n = 4). The experiment was performed 3 times with similar results. (E) C57BL/6 mice received anti–IL-1β antibody or vehicle 24h before TBI or control treatment. RNA from the small intestine was isolated 40 h after TBI, and the expression of IL-1β was determined by quantitative PCR (qPCR). Each data point represents an individual animal (n = 4). The experiment was performed three times, with similar results. (F) BALB/c mice underwent different conditioning regimes: TBI alone (day 0; n = 3) or followed by allo-HCT with C57/BL6 BM plus T cells (day 10; n = 4); BU/CY chemotherapy or FLU/CY chemotherapy alone (day 0, n = 5) or followed by allo-HCT with C57/BL6 BM plus T cells (day 3, n = 6) as indicated. Untreated mice (n = 9) served as control. At the indicated time points, peritoneal fluid was isolated and UA levels were determined. Each data point represents an individual animal, and the experiment was performed twice with similar results. (G) BALB/c mice underwent TBI followed by allo-HCT with C57/BL6 BM alone (n = 10) or C57/BL6 BM plus T cells. Mice were treated with PBS (n = 16) or uricase starting from day −1 forward or from day +5 forward (n = 10 per group), as indicated. The experiment was performed twice and the resulting survival data were pooled. (H) On day 3 after allo-HCT, serum was isolated from BALB/c mice that received treatment with PBS or uricase from day 0 forward. Serum IL-1β levels were determined by ELISA. ***, P = 0.0001; **, P < 0.01; *, P < 0.05.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal