Objective:To comparison of three different beta blockers on murine hemangioma (EOMA cells) cells in vitro and in vivo effects.Preliminary study on the therapeutic effect of propranolol on vascular tumor in mice and possible mechanisms , provide a reference for beta blockers in the treatment of infantile hemangioma .Methods: Comparative study on the effects of three kinds of different β-receptor blockers---metoprolol, propranolol and butoxamine , on the proliferation and apoptosis of Mouse Hemangioendothelioma Endothelial cell (EOMA cells) was conducted in vitro.EOMA cells were cultured in vitro,randomly divided into different groups,propranolol and timolol were added into the medium respectively ,after 24 h intervention.MTT assay and acridine orange staining assay were conducted respectively to detect cell viability and apoptosis level .EOMA cells were transplanted into nude mice in vivo.Tumor volume growth to 100 mm3 ,animals were randomly divided into 4 groups respectively ,the control group ,metoprolol group,Bhutto Samin group and propranolol group ,drug group according to 2 mg/( kg? d) oral gavage ,control group were given an equal volume of saline ( NS ) , every two days measurement tumor volume size .Serum levels of tumor necrosis factor alpha ( TNF-α) and vascular endothelial growth factor ( VEGF ) were detected by enzyme linked immunosorbent assay ( ELISA ) in the end of the experiment.Results:For propranolol,after 24 h treatment,significant differences of cell viability and apoptosis were noted (P<0.05) at the concentration of 50 μmol/L,while continuing to increase to 800 μmol/L,the cell survival rate decreased sharply to close to 10%. Acridine orange staining at the 50 μmol/L group after 24 h revealed many apoptotic cells .For metoprolol and butoxa mine ,significant differences of cell viability and apoptosis were noted ( P<0.05 ) at the concentration of 100 μmol/L,while continuing to increase to 800μmol/L,the cell survival rate decreased sharply to close to 20%.It was significantly higher than propranolol group at the same concentration ( P<0.05 ) .It showed a similar trend in acridine orange staining .In vivo experiments showed that the end of the experiment of metoprolol , butoxamine group and propranolol drugs in mice tumor volume , respectively ( 1 642.8 ±89.3 ) , ( 1 529.3 ± 119.1) and (752.7±46.5)mm3,significantly lower than the control group of mice tumor volume of (2 023.3±123.0) mm3(P<0.001).Metoprolol,butoxamine mice and propranolol drugs group ,serum VEGF levels for (606.5±105.8 ) pg/ml,(534.3±243.2 ) pg/ml and (420.1±123.7) pg/ml, significantly lower than the PBS control group [(825.8±145.7) pg/ml,(P<0.05)],the TNF alpha result was followed by(301.3±62.3) pg/ml,(305.1±53.8) pg/ml and (288.8±59.5) pg/ml,significantly lower than the normal control group [(444±100.4) pg/ml,P<0.05].Conclusion:Three kinds of beta-blockers can effectively inhibit EOMA cells proliferation and induce apoptosis in vitro, the role of propranolol more significantly than butoxamine and metoprolol .Three kinds of beta blockers restrain the growth of the hemangioma in vivo ,in which the inhibitory effect of propranolol is stronger than the metoprolol and butoxa mine.Three kinds of beta blockers can lower the levels of VEGF and TNF-αin vivo.Indicating that propranolol on vascular tumor in mice may be one of the mechanisms of β1 and β2 receptor synergy effect and its mechanism in the treatment of hemangioma may be associated with VEGF and TNF-α.

OBJECTIVE To analyze trimethylation of genome-wide histone H3 lysine 4(H3K4met3) induced by silicon dioxide(SiO2)through chromatin immunoprecipitation linked to microarrays(ChIP-chip)in lung fibroblast(LF)of rats. METHODS A primary co-culture model of rat alveolar macrophages (AM)and LF in vitro. AM were exposed to 100 mg · L-1 free SiO2 for 24 h,before LF were collected and the phenotype of LF was determined after transdifferentiation by immunohistochemistry. ChIP-chip was used to profile the variations of trimethylation in H3K4 of lung fibroblasts in CpG island regions. ChIP-qPCR was used to validate the microarray results. The mRNA expression of nfib and kpna3 was analyzed by qRT-PCR. RESULTS Totally 1815 (518 increased and 1297 decreased) genes of H3K4met3 displayed significant differences in SiO2 100 mg·L-1 group compared with control group(Cy3/Cy5 value>2.0 or <0.5,NimbleScan V2.5 software). The results of ChIP-qPCR were quite consistent with those of microarray. CONCLUSION There are significant differences in methylation of genome-wide H3K4 between SiO2 100 mg·L-1 group and control group. These novel candidate genes may become potential biomarkers or new interfered targets.

Objective To investigate the value of non-contrast CT(NCCT)-based radiomics for identifying acute unilateral middle cerebral artery occlusion(MCAO)with negative hyperdense artery sign(HAS).Methods All 80 patients with acute unilateral MCAO confirmed by angiography(MR angiography[MRA]or CT angiography[CTA]or DSA)and presenting with negative NCCT presentation for HAS were enrolled from January 2015 to June 2023 in the Emergency Department of Stroke Center of Affiliated Hospital of Yangzhou university.On the NCCT images,the occluded segment of the middle cerebral artery on the affected side of each case and the corresponding segment of the vessel on the normal side were used as the regions of interest,and a total of 108 radiomic features were extracted.The least absolute shrinkage and selection operator(LASSO)was used to screen the key features,construct and calculate the radiomics score,and four imaging histology models,support vector machine(SVM),light gradient boosting machine(LightGBM),GradientBoosting and adaptive boosting(AdaBoost),were built respectively to predict MCAO.Predictive performance was evaluated by the area under the receiver operating characteristic curves,and comparisons between the modeled receiver operating characteristic curves were made using the Delong test.Finally,the value of the application of radiological modeling was assessed by clinical decision curve analysis(DCA).Results The NCCT images based on 160 vessels were finally screened for 6 key features,including skewness,energy,gray level size zone matrix(GLSZM)-gray uneven,GLSZM-low gray area emphasis,GLSZM-size area non-uniform standardization,GLSZM-area entropy.The area under the curve(AUC)of the SVM-test was 0.688(95％CI 0.497-0.878)with an accuracy of 0.688;the AUC of the LightGBM-test was 0.787(95％CI 0.620-0.955)with an accuracy of 0.781;the AUC of the GradientBoosting-test was 0.654(95％CI 0.457-0.852)with an accuracy of 0.688;the AUC of the AdaBoost-test was 0.707(95％CI 0.515-0.899)with an accuracy of 0.750.The Delong test showed a statistically significant difference between LightGBM-test and GradientBoosting-test(P=0.040),and no statistically significant difference in performance between the remaining models(all P＞0.05).DCA showed that the LightGBM-test performed better.Conclusion NCCT-based radiomics has good diagnostic efficacy for identifying acute unilateral MCAO with negative HAS,and this conclusion needs to be further verified by multi-center and large sample studies.

Objective:To study the role and specific mechanisms of glucose-6-phosphate dehydrogenase (G6PD), a key enzyme for glycometabolism, mediated reduction stress in arsenic-induced malignant transformation of cells.Methods:Immortalized human skin keratinocyte-forming cells (HaCaT cells) were treated with sodium arsenite (NaAsO 2) at a concentration of 0.0 (control group) and 1.0 μmol/L (arsenic group), and malignant transformation indicators were tested using cell growth kinetics assay, cell scratch assay, and soft agar colony formation assay. At different stages of arsenic treatment (0, 1, 7, 14, 21, 28, 35 passages of cells), the effects of NaAsO 2 on glycometabolism in HaCaT cells were determined using corresponding reagent kits and Western blot, including glucose-6-phosphate (G6P), lactate, acetyl CoA, G6PD levels, as well as protein expression levels of hexokinase 2 (HK-2), 6-phosphofructose-2-kinase/fructose-2, 6-diphosphatase 3 (PFKFB3), pyruvate dehydrogenase kinase 1 (PDK1), 6-phosphoglucose dehydrogenase (PGD), and G6PD. Mitochondria were extracted, and the effects of NaAsO 2 on HaCaT cells and mitochondrial redox [reduced nicotinamide adenine dinucleotide phosphate (NADPH)/nicotinamide adenine dinucleotide phosphate (NADP +) ratio, reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio] were determined using corresponding reagent kits. The effect of G6PD on reduction stress and NaAsO 2-induced malignant transformation of HaCaT cells was determined using small interfering RNA (siRNA) intervention method. Results:Compared 1.0 μmol/L NaAsO 2-cultured HaCaT cells up to 35 generations (T-HaCaT cells) with matching passage 0.0 μmol/L NaAsO 2-cultured HaCaT cells [(33.797 ± 0.280) h, 0.177 ± 0.015, 13.667 ± 2.625], the multiplication time [(24.042 ± 0.479) h] was shorter ( t = 30.45, P < 0.001), the cell migration rate (0.396 ± 0.039) was higher ( t = 9.08, P < 0.001), and the number of colonies formed in soft agar (73.667 ± 4.450) was higher ( t = 20.11, P < 0.001). Compared with matching passage control group cells and 0 generation of the same group, G6P level in the arsenic group was higher at passages 1, 7, 14, 21, 28 and 35 ( P < 0.05), lactate and G6PD levels were higher at passages 14, 21, 28 and 35 ( P < 0.05), acetyl CoA level was lower at passages 21, 28 and 35 ( P < 0.05), and protein expression levels of HK-2, PFKFB3, PDK1, PGD, and G6PD were higher at passages 7, 14, 21, 28 and 35 ( P < 0.05). The NADPH/NADP + ratio of cells was higher at passages 1, 14, 21, 28 and 35 ( P < 0.05), GSH/GSSG ratio was higher at passages 21, 28 and 35 ( P < 0.05). The ratio of mitochondrial NADPH/NADP + was higher at passages 1, 7, 21, 28 and 35 ( P < 0.05), the GSH/GSSG ratio was higher at passages 1, 7, 14, 21, 28 and 35 ( P < 0.05). G6PD expression was silenced by siRNA in T-HaCaT cells, compared with the T-HaCaT con siRNA-treated group, the T-HaCaT G6PD siRNA-treated group had lower NADPH/NADP + and GSH/GSSG ratios in both cells and mitochondria ( P < 0.05), longer cell multiplication time, lower cell migration rate, and fewer soft agar colonies ( P < 0.05). Conclusion:During the malignant transformation of HaCaT cells induced by NaAsO 2, G6PD and other enzymes related to glycometabolism are activated, leading to reprogramming of glycometabolism, resulting in an imbalance of cell redox homeostasis and enhanced reduction stress in cells and mitochondria, thereby promoting NaAsO 2 induced malignant transformation of HaCaT cells.

OBJECTIVE: To investigate the epithelial-mesenchymal transition(EMT) induced by direct or indirect exposure to free silicon dioxide(SiO_2) and the expression of surface protein marker in rat typeⅡalveolar epithelial cell RLE-6TN.METHODS: i) The alveolar macrophages(AM) were isolated from specific pathogen-free SD rat by bronchoalveolar lavage.AM and RLE-6TN were treated with 0-140 mg/L(final concentration) of SiO_2 suspension and were cultured conventionally for 24,48 and 72 hours. The cell viability was detected by CCK-8 assay. The result of CCK-8 essay was used to choose the SiO_2 concentration for the following study. ii) To establish models of RLE-6TN co-cultured with AM that were seeded in Transwell. The cells were divided into 4 groups: the direct control group(RLE-6TN,no SiO_2 exposed),the direct exposure group(RLE-6TN,treated with 100 mg/L SiO_2),the indirect control group(RLE-6TN and AM were cocultured,no SiO_2 exposed) and the indirect exposure group(RLE-6TN and AM were co-cultured,AM was treated with 100 mg/L SiO_2 directly). Western blotting was used to detect the expression of E-cadherin(E-cad) and α-smooth muscle protein(α-SMA) after cells were cultured for 0,24,48 and 72 hours. RESULTS: i) According to the CCK-8 assay,the final concentration of 100 mg/L SiO_2 was chosen for the following study. ii) The difference of relative expression of E-cad andα-SMA in RLE-6TN was statistically significant in different treatment combination and time(P < 0. 01). The E-cad expression of RLE-6TN at 48 and 72 hours in the direct exposure group and the indirect exposure group was lower than that in direct control group at the same time point(P < 0. 05). The E-cad expression in RLE-6TN at 72 hours in the direct exposure group was lower than that in the 0 and 24 hours(P < 0. 05). The E-cad expression in RLE-6TN at 48 and 72 hours in the indirect exposure group was lower than that in the 0 hour(P < 0. 05). At 48 and 72 hours,the α-SMA expression in the indirect exposure group and the direct exposure group was higher than that in their control groups at the same time point(P < 0. 05). The expression of α-SMA in the indirect exposure group was higher than that in the direct exposure group(P < 0. 05). The expression of α-SMA in both exposure groups increased in a time-effect relationship(P <0. 05). CONCLUSION: Direct or indirect exposure to free SiO_2 can induce EMT in RLE-6TN,and decrease the expression of E-cad and increase the expression of α-SMA in a time-effect relationship. Indirect exposure group is more susceptible to EMT.