AIM:To establish an optimal protocol for isolating and extracting mouse hepatocellular carcinoma H22 cell-detrived microparticles(H22-MPs),evaluate their stimulating effect on dendritic cells(DC),and investigate the cytotoxic capacity of CD8+T cells induced by H22-MPs on hepatocellular carcinoma cells.METHODS:The H22-MPs were extracted by high-speed differential centrifugation,and the morphology of H22-MPs was observed by transmis-sion electron microscopy.The particle size distribution of H22-MPs was detected by dynamic light scattering,and the ex-pression of vesicle surface markers was detected by Western blot.The protein components and pathways involved by MPs were analyzed by labeling free quantitative proteomics combining with bioinformatics GO and KEGG enrichment methods.The effect of MPs antigen on the expression of CD11c,a mature marker of DC,was detected by laser confocal microscopy.Next,CD8+T cells were sorted by magnetic beads,and the secretion of tumor necrosis factor-α(TNF-α),interferon-γ(IFN-γ)and interleukin-2(IL-2)in the supernatant of CD8+T cells stimulated by DC loaded with H22-MPs was detected by ELISA.Lactate dehydrogenase(LDH)test was conducted to evaluate the specific killing effect of CD8+T cells induced by DC loaded with H22-MPs on hepatocellular carcinoma cells.RESULTS:The results showed that the morphology,size and vesicle markers of H22-MPs met the requirements.Most of the H22-MPs protein antigens were derived from its parent cell H22 and coordinated various signal transduction pathways in the cells.Subsequently,it was detected that H22-MPs increased the expression of CD11c on the surface of DC,and H22-MPs were mainly localized in the lysosomes of DC.In addition,DC loaded with H22-MPs stimulated CD8+T to release high levels of TNF-α,IFN-γ and IL-2,and it also pro-moted CD8+T to overexpress CD69,thus inducing CD8+T to killhepatocellular carcinoma cells specifically.CONCLU-SION:H22-MPs promote the maturation of DC,and the mature DC may induce CD8+T cells to play an anti-hepatocellu-lar carcinoma immune response by promoting the activation of CD8+T cells.

AIM:To establish an optimal protocol for isolating and extracting mouse hepatocellular carcinoma H22 cell-detrived microparticles(H22-MPs),evaluate their stimulating effect on dendritic cells(DC),and investigate the cytotoxic capacity of CD8+T cells induced by H22-MPs on hepatocellular carcinoma cells.METHODS:The H22-MPs were extracted by high-speed differential centrifugation,and the morphology of H22-MPs was observed by transmis-sion electron microscopy.The particle size distribution of H22-MPs was detected by dynamic light scattering,and the ex-pression of vesicle surface markers was detected by Western blot.The protein components and pathways involved by MPs were analyzed by labeling free quantitative proteomics combining with bioinformatics GO and KEGG enrichment methods.The effect of MPs antigen on the expression of CD11c,a mature marker of DC,was detected by laser confocal microscopy.Next,CD8+T cells were sorted by magnetic beads,and the secretion of tumor necrosis factor-α(TNF-α),interferon-γ(IFN-γ)and interleukin-2(IL-2)in the supernatant of CD8+T cells stimulated by DC loaded with H22-MPs was detected by ELISA.Lactate dehydrogenase(LDH)test was conducted to evaluate the specific killing effect of CD8+T cells induced by DC loaded with H22-MPs on hepatocellular carcinoma cells.RESULTS:The results showed that the morphology,size and vesicle markers of H22-MPs met the requirements.Most of the H22-MPs protein antigens were derived from its parent cell H22 and coordinated various signal transduction pathways in the cells.Subsequently,it was detected that H22-MPs increased the expression of CD11c on the surface of DC,and H22-MPs were mainly localized in the lysosomes of DC.In addition,DC loaded with H22-MPs stimulated CD8+T to release high levels of TNF-α,IFN-γ and IL-2,and it also pro-moted CD8+T to overexpress CD69,thus inducing CD8+T to killhepatocellular carcinoma cells specifically.CONCLU-SION:H22-MPs promote the maturation of DC,and the mature DC may induce CD8+T cells to play an anti-hepatocellu-lar carcinoma immune response by promoting the activation of CD8+T cells.

Objectives To study the effect of cadmium (Cd) on the proliferation and apoptosis of mouse spermatocyte (GC-2 spd) cells and explore the underlying molecular mechanism. Methods GC-2 spd cells were cultured with 0, 5, 10, 15, 20 and 30 μM CdCl2, respectively, for 24 hours. The cell viability and IC50 of Cd were estimated based on CCK-8 data. The apoptosis of GC-2 spd cells and cellular concentration of ROS were analyzed by flow cytometry after treatment of the cells with different concentrations of CdCl2 (0, 5, 10 μM) for 24 hours. The expression levels of JNK/c-Jun signaling pathway regulatory proteins, pro-apoptotic factor Bax and anti-apoptotic factor Bcl-2, were determined by Western blot. Results Cd inhibited the proliferation of GC-2 spd cells with IC50 value of 12.99 μM, 95% CI [11.95, 14.00]. Exposure to 5 and 10 μM CaCl2 resulted in increases in apoptosis and cellular ROS generation in a dose-dependent manner, which was statistically significant compared with the control (P < 0.05). Although there was no difference in the expression level of c-Jun (P > 0.05), the phosphorylation level of JNK and c-Jun in Cd group was highly increased as compared to the control (P < 0.05). In addition, Cd exposure significantly increased the expression of Bax protein but decreased the expression Bcl-2 protein (P < 0.05). Conclusions Cadmium induces GC-2 spd cell apoptosis by increasing concentration of ROS and regulating the JNK/c-Jun signaling pathway.

OBJECTIVETo observe the effects of conventional western medication and joss stick moxibustion combined with pricking and cupping for herpes zoster in acute stage, and to explore its analgesic mechanism.

METHODSSeventy patients with acute herpes zoster were randomized into an observation group (33 cases after 2 dropping) and a control group (34 cases after 1 dropping). Patients in the observation group were treated with joss stick moxibustion combined with pricking and cupping at localpoints for 7 times, once every other day. Oral acyclovir, vitamin Band mecobalamin tablets were applied in the control group for continuous 14 days, and interferon injection was used for continuous 6 days, etc. The herpes evaluation indexes of blister stopping time, scab time and decrustation time as well as pain intensity were observed before and after treatment. Peripheral serum substance P (SP) content of herpes local situation was detected. The comprehensive effects were evaluated.

RESULTSThe blister stopping time, scab time and decrustation time in the observation group were shorter than those in the control group (all<0.05). There was no statistical significance for pain relief degree between the two groups (>0.05). The pain beginning to ease time and duration time in the observation group were better than those in the control group (both<0.05). The contents of SP in the two groups decreased after treatment (both<0.01), and it was better in the observation group (<0.05). The total effective rate of the observation group after treatment was 87.9% (29/33), and that of the control group was 85.3% (29/34), which were not statistically significant (>0.05). The cured rate of the observation group was better than that of the control group [66.7% (22/33) vs 58.8% (20/34),<0.05].

CONCLUSIONJoss stick moxibustion combined with pricking and cupping are effective for herpes zoster, which have quicker and good analgesic effects than conventional western medication. Its mechanism may be related to reducing the content of SP more fast and to a larger degree.

Objective To investigate the design and manufacture of 3D printed compensator in electron radiation therapy for Merkel cell carcinoma,and to verify the feasibility of this technique in electron radiation therapy.Methods Computed tomography was used to collect images of a human head phantom.The delineation of target volume of Merkel cell carcinoma was simulated in the planning system and a radiotherapy plan was formulated after adding the compensator.The compensator was printed out by a 3D printer and fixed on the head phantom.A second CT scan was performed to make a new treatment plan.For the two plans,several planes parallel to the beam were selected to calculate gamma passing rates.The actual dose distribution was measured using disposable films.The gamma passing rate was compared between the film system and the planning system.The conformity index (CI) and the heterogeneity index (HI) of target volume were compared between the plans using the printed compensator and the conventional compensator of the same thickness.Comparison between the two plans was made by paired t test.Results Using the dose distribution of the plan with simulated compensator,the gamma passing rate was 94.7±2.3％ in the plan with 3D printed compensator.Using the dose distribution measured by the film,the gamma passing rate was 96.6％ in the plan with 3D printed compensator.Compared with the conventional compensator,the 3D printed compensator achieved a significantly elevated CI (0.85 vs.0.69,P=0.004) and a slightly improved HI (1.30 vs.1.26,P=0.001).Conclusions The conformal dose distribution provided by 3D printed compensator for tumors at different depths meets the clinical need.