Objective To investigate the effect of peptidoglycan (PGN) on the secretion of pro-inflammatory cytokines by dendritic cells (DCs) and the regulation of T helper 17 (Th17) responses in experimental autoimmune uveitis. Methods Bone marrow cells from naive mice were cultured with granulocyte macrophage-colony-stimulating factor and interleukin (IL)-4 to induce DCs. DCs cultured for six days were randomly divided into two groups: PGNtreated group and control group. The DCs in PGNtreated group were stimulated with PGN and the same volume of phosphate buffered saline was added to the DCs as control group. The relative mRNA expression levels of IL-23, tumor necrotic factor alpha; (TNF-alpha;), IL-6,IL-1beta;were measured by real-time reverse transcriptase polymerase chain reaction (RT-PCR). Peptide fragment of interphotoreceptor retinoidbinding protein (IRBP1-20)specific T cells, which were isolated from the spleen and draining lymph nodes of C57BL/6 mice immunized with IRBP1-20 peptide fragments 13 days earlier, were co-cultured with PGN-treated or untreated DCs, respectively. Total RNA from T cells cocultured for two days were isolated and the relative expression of retinoic acid receptor-related orphan receptor gamma;t (ROR-gamma;t), IL-17, T-box expression in T cells (T-bet), interferon gamma; (IFN-gamma;) mRNA were detected by realtime RT-PCR. On the second, the fifth and the seventh day, the cocultured T cells were analyzed by flow cytometry to detect the percentages of IFN-gamma;, IL-17 positive cells. Results The real-time RT-PCR results revealed that the level of IL-23, IL-1beta;, IL-6, TNF-alpha; mRNA from PGNstimulated DCs were significantly increased compared to the control group (t=-14.363, -5.627, -3.85, -28.151; P＜0.05). The level of RORgamma;t, IL-17 mRNA from the T cells cocultured with PGN-stimulated DCs were greatly increased compared with the control group (t=-5.601, -19.76；P＜0.05). However, the level of T-bet, IFN-gamma; mRNA from the T cells cocultured with PGNstimulated DCs were significantly decreased compared with the control group (t=4.717, 11.207; P＜0.05). Data of flow cytometry showed that at two days, five days, seven days after cocultured with PGN-treated DCs, the percentages of IL-17 positive T cells were increased compared to the control group (t=-2.944, -3.03, -4.81; P＜0.05), and the percentages of IFN-gamma; positive T cells had no remarkable change (t=-1.25, -0.18, -2.16; P＞0.05). Conclusion PGN can promote the secretion of Th17-related cytokines by DCs, which favors proliferation and differentiation of Th17 in experimental autoimmune uveitis.
Patient-specific volumetric modulated arc therapy (VMAT) quality assurance (QA) process is an important component of the implementation process of clinical radiotherapy. The tolerance limit and action limit of discrepancies between the calculated dose and the delivered radiation dose are the key parts of the VMAT QA processes as recognized by the AAPM TG-218 report, however, there is no unified standard for these two values among radiotherapy centers. In this study, based on the operational recommendations given in the AAPM TG-218 report, treatment site-specific tolerance limits and action limits of gamma pass rate in VMAT QA processes when using ArcCHECK for dose verification were established by statistical process control (SPC) methodology. The tolerance limit and action limit were calculated based on the first 25 in-control VMAT QA for each site. The individual control charts were drawn to continuously monitor the VMAT QA process with 287 VMAT plans and analyze the causes of VMAT QA out of control. The tolerance limits for brain, head and neck, abdomen and pelvic VMAT QA processes were 94.56%, 94.68%, 94.34%, and 92.97%, respectively, and the action limits were 93.82%, 92.54%, 93.23%, and 90.29%, respectively. Except for pelvic, the tolerance limits for the brain, head and neck, and abdomen were close to the universal tolerance limit of TG-218 (95%), and the action limits for all sites were higher than the universal action limit of TG-218 (90%). The out-of-control VMAT QAs were detected by the individual control chart, including one case of head and neck, two of the abdomen and two of the pelvic site. Four of them were affected by the setup error, and one was affected by the calibration of ArcCHECK. The results show that the SPC methodology can effectively monitor the IMRT/VMAT QA processes. Setting treatment site-specific tolerance limits is helpful to investigate the cause of out-of-control VMAT QA.