However, ROS levels in the thymus were elevated. was elevated in the tumors and thymuses of the HBO group. Conclusion HBO induced ROS signaling in the thymus, inhibited CD3+ T cell generation, and facilitated malignant glioma cell growth in the intracranial glioma mouse model. bioluminescence imaging (BLI) 10 days after the intracranial transplantation of GL261-Luc glioma cells. In accordance with the manufacturers protocol, individual mice were put into the chamber of an ICE Chemi & Fluo System P80 BLI system (Photometrics; Tucson, AZ, USA). The exposure time was set to 8 min. The tumor size was quantified with normalized photon ?ux. Hyperbaric oxygen intervention process Mice of the experimental group were placed into NG90-IIIB medical hyperbaric oxygen chambers (Ningbo Hyperbaric Oxygen Chamber Manufacturing plant, Ningbo, China), and underwent standard HBO intervention according to the manufacturers Amoxicillin trihydrate protocol (2.5 atmospheres, 0.015 MPa/minute for 10 minutes; maintain for 60 moments and decompress at the same rate). HBO treatment was started 10 days after tumor cell injection, and was performed daily for 10 days. All applicable international, national, and institutional guidelines for the care and use of animals were followed. The complete study protocol was approved by the Ethics Committee of the First Affiliated Hospital of Kunming Medical University or college. Small animal magnetic resonance imaging MRI of mouse brains was performed 20 days after glioma cell injection (10 days after HBO Amoxicillin trihydrate treatment). Mice were anesthetized (1% isoflurane Amoxicillin trihydrate treatment, and intraperitoneal injection of 0.8?mL/kg gadopentetate dimeglumine), then imaged with an 8.5-T Biospec vertical bore system (Bruker, Billerica, MA, USA). MRI images were generated and the tumor volume (V) was measured using a threshold method according to an evaluation of length (L), width (W), and height (H), using the formula: V?=?(L??W??H)/2. Circulation cytometry assays The ROS level was evaluated by circulation cytometry using the DCFDA Cellular Reactive Oxygen Species Detection Assay Kit Rabbit polyclonal to EPHA4 (Abcam, Cambridge, MA, USA) according to the manufacturers protocol. Briefly, cells were stained with 2,7-dichlorofluorescin diacetate (DCFA) at 37C for 30 minutes, washed with 1 buffer, and the transmission was go through at an excitation of 485 nm and an emission of 535 nm. The expression of markers (including CD3, CD4, CD8, CD25, and FoxP3) on T cells from different organs was determined by circulation cytometry as previously explained.20 Statistical analysis In comparisons of the HBO group and the control group, values were calculated by two-tailed Students by BLI 10 days after GL261-Luc glioma cells intracranial transplantation. Tumor growth rates were similar between the HBO experimental group and the control group (Photon area pixels: 5678957 vs. 60691400, respectively; Photon sum 1000: 498.8111.8 vs. 476.777.7, respectively). However, 10 days after HBO treatment, the HBO group showed significantly larger tumors than the control group (Photon area pixels: 222207780 vs. 456407191, respectively, (left panel: 10 days after transplantation; middle panel: 20 days after transplantation, control group; right panel: 20 days after transplantation, HBO group). (b) Photon sum of the HBO group and the control group. (c) Photon light-emitting area of the HBO group and the control group. (d) Representative magnetic resonance images of tumors inoculated in the HBO group and the control group (left panel: control group; right panel: HBO group). (e) Predicted tumor sizes of tumors inoculated in the HBO group and the control group. HBO, hyperbaric oxygen. To further confirm the effects of HBO on intracranial glioma cell growth, MRI was performed 20 days after the.