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OBJECTIVE@#To investigate the role and molecular mechanism of exosomal miR-224-5p in colorectal cancer (CRC).@*METHODS@#The miR-224-5p expression in CRC patient tissues and cell-derived exosomes was measured by laser capture microdissection and qRT-PCR, respectively. Dual-luciferase reporter gene assay was used to determine the target gene of miR-224-5p. The protein expressions of p53 and unc-51 like kinase 2 (ULK2) in CRC cells were detected by western blot. Flow cytometry was used to detect cell cycle and apoptosis. Cell proliferation was measured by CCK8 and EdU assay.@*RESULTS@#The miR-224-5p expression was upregulated in CRC tissues and increased progressively with the rise of CRC stage. CRC cells secreted extracellular miR-224-5p mainly in an exosome-dependent manner, and then miR-224-5p could be transferred to surrounding tumor cells to regulate cell proliferation in the form of autocrine or paracrine. Moreover, ULK2 was characterized as a direct target of miR-224-5p and was downregulated in CRC tissues. Interestingly, ULK2 inhibited CRC cell proliferation in a p53-dependent manner. Furthermore, exosome-derived miR-224-5p partially reversed the proliferation regulation of ULK2 on CRC cells.@*CONCLUSION@#Our findings demonstrate that exosome-transmitted miR-224-5p promotes p53-dependent cell proliferation by targeting ULK2 in CRC, which may offer promising targets for CRC prevention and therapy.
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Humans , MicroRNAs/metabolism , Tumor Suppressor Protein p53/metabolism , Exosomes/metabolism , Cell Proliferation/genetics , Colorectal Neoplasms/genetics , Cell Line, Tumor , Gene Expression Regulation, NeoplasticABSTRACT
Objective: To investigate the metabolic stability, the main CYP450 enzymes phenotypes and metabolites of Diosbulbin B based on in vitro metabolism model. Methods: For metabolic stability study, UPLC-MS/MS was used to detect the remaining Diosbulbin B content in the incubation solution after being incubated with human and rat liver microsomes, respectively. Ten recombinant human CYP450 enzymes (1A1, 1A2, 1B1, 2A13, 2A6, 2B6, 2D6, 2C9, 2C19, 3A4) were used for identifying the metabolic enzyme phenotypes of Diosbulbin B. Moreover, the major metabolic enzyme phenotype for the metabolism of Diosbulbin B was confirmed and verified by the rat isolated hepatic perfusion model. The metabolites of Diosbulbin B in human and rat liver microsomes were determined by LC-MS/MS. Results: The metabolic percentage of Diosbulbin B in human and rat liver microsomes were 37% and 59%, respectively. Its half-lives t1/2 in human and rat liver microsomes were 97.4 and 52.3 min, respectively. The intrinsic clearance rates CLint in human and rat livers were 8.23 and 23.9 mL/(min•kg), and liver clearance CLh in human and rat livers were 5.89 and 16.8 mL/(min•kg). It can be found that the metabolic rate of Diosbulbin B in rat liver microsomes was faster than in human liver microsomes. There were five CYP enzymes, including 3A4, 2C19, 2C9, 1A13 and 1A1, related to the metabolism of Diosbulbin B, especially CYP3A4. The hepatic perfusion experimental results showed that the metabolism of Diosbulbin B was inhibited by ketoconazole, and the inhibitory effect was enhanced along with the increasing dosage of ketoconazole, which confirmed that CYP3A4 played an important role in metabolism of Diosbulbin B. There was one metabolite (M1) of Diosbulbin B has been found in both human and rat liver microsomes incubation. Conclusion: The metabolic rate of Diosbulbin B in rat liver microsomes was faster than human liver microsomes. The CYP3A4 plays a leading role in the metabolism of Diosbulbin B. And a demethylated metabolite of Diosbulbin B was appeared in both human and rat liver microsomes incubation.
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Objective To explore the working mode of continuous malaria elimination. Methods Judong Village in Congjiang County, Guizhou Province, which had the highest record of malaria incidence in the history of Guizhou Province, was selected as a pilot, a comprehensive prevention and control intervention was carried out aiming at the malaria epidemic, transmission and influencing factors, and the effect was evaluated after the intervention. Results From 2015 to 2016, through those intensive measures of constructing system, improving environment and health education, the people’s awareness of malaria prevention was increased, the people’s enthusiasm and initiatives to participate in the prevention and control of malaria were improved in Judong Village. Meanwhile, the roads were hardened and beautified, the dispose of sewage, feces and garbage was centralized. The awareness rate of malaria prevention knowledge of residents increased to 85% to 95%, and the formation rate of anti-mosquito behavior increased to 89.66%. The density of media Anopheles reduced from 0.56 to 0.07 mosquitoes per hour each mosquito catching platform. Conclusion A long-term mechanism for malaria prevention and control has been almost constructed in Judong Village, which facilitates the consolidation of malaria control achievements, and the progress towards malaria elimination.
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Objective To explore the working mode of continuous malaria elimination. Methods Judong Village in Congjiang County, Guizhou Province, which had the highest record of malaria incidence in the history of Guizhou Province, was selected as a pilot, a comprehensive prevention and control intervention was carried out aiming at the malaria epidemic, transmission and influencing factors, and the effect was evaluated after the intervention. Results From 2015 to 2016, through those intensive measures of constructing system, improving environment and health education, the people’s awareness of malaria prevention was increased, the people’s enthusiasm and initiatives to participate in the prevention and control of malaria were improved in Judong Village. Meanwhile, the roads were hardened and beautified, the dispose of sewage, feces and garbage was centralized. The awareness rate of malaria prevention knowledge of residents increased to 85% to 95%, and the formation rate of anti-mosquito behavior increased to 89.66%. The density of media Anopheles reduced from 0.56 to 0.07 mosquitoes per hour each mosquito catching platform. Conclusion A long-term mechanism for malaria prevention and control has been almost constructed in Judong Village, which facilitates the consolidation of malaria control achievements, and the progress towards malaria elimination.
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Objective To establish an HPLC method to determine the related substances of metolazone and valsartan in com?pound metolazone tablets. Methods An Agilent Eclipse SB-C18 column (4.6 mm × 250 mm,5 μm) was used with 0.01 mol/L KH2PO4 buffer(pH=3.5)-acetonitrile as the mobile phase with gradient elution at a flow rate of 1.0 ml/min. The column temperature was 30℃ and the detection wavelength was 237 nm. Injection volume was 20 μl. Results Metolazone,valsartan and related sub?stance B of valsartan were separated completely. The calibration curves were linear within the range of 3-30μg/ml for metolazone, 0.1-2.0μg/ml for valsartan and 0.08-2.0μg/ml for related substane B of valsartan. The average recoveries were 102.97%,100.81%and 100.44%,respectively. The repeatability and intermediate precision met with requirements. The test solution was stable within 24 h. Conclusion The method is specific,sensitive,accurate and reliable,thereby can be used for the determination of metolazone and valsartan related substances in compound metolazone tablets.
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Objective To establish an in vitro accelerated release method of triptorelin acetate microspheres with good in vi-tro/in vivo correlation(IVIVC). Methods The in vivo release of triptorelin acetate from microspheres was obtained by residual method in rats. Influences of pH value,concentration of ethanol,temperature,rotation speed and concentration of antiseptic on the in vitro accel-erated release were studied,then the correlation between in vitro accelerated release and in vivo release of the microspheres was estab-lished by adjusting the release conditions. Results The in vitro accelerated release medium of triptorelin acetate microspheres composed of 15%ethanol solution(containing 0.06%Tween 80 and 0.1%benzalkonium chloride)at 55℃with rotating rate of 200 r/min. The cumulative release of triptorelin acetate from microspheres was 87.35%at 30 h under accelerated release condition,equivalent to in vivo release for 30 days. The established in vitro accelerated release had a good correlation with that of in vivo(y=0.8845x+12.4510, R2=0.9938). Conclusion The in vitro accelerated release of triptorelin acetate microspheres could correlate well with in vivo release and has a potential application in rapid and effective evaluation of triptorelin acetate microspheres.
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OBJECTIVES: Recent guidelines recommend that all cirrhotic patients should undergo endoscopic screening for esophageal varices. That identifying cirrhotic patients with esophageal varices by noninvasive predictors would allow for the restriction of the performance of endoscopy to patients with a high risk of having varices. This study aimed to develop a decision model based on classification and regression tree analysis for the prediction of large esophageal varices in cirrhotic patients. METHODS: 309 cirrhotic patients (training sample, 187 patients; test sample 122 patients) were included. Within the training sample, the classification and regression tree analysis was used to identify predictors and prediction model of large esophageal varices. The prediction model was then further evaluated in the test sample and different Child-Pugh classes. RESULTS: The prevalence of large esophageal varices in cirrhotic patients was 50.8 percent. A tree model that was consisted of spleen width, portal vein diameter and prothrombin time was developed by classification and regression tree analysis achieved a diagnostic accuracy of 84 percent for prediction of large esophageal varices. When reconstructed into two groups, the rate of varices was 83.2 percent for high-risk group and 15.2 percent for low-risk group. Accuracy of the tree model was maintained in the test sample and different Child-Pugh classes. CONCLUSIONS: A decision tree model that consists of spleen width, portal vein diameter and prothrombin time may be useful for prediction of large esophageal varices in cirrhotic patients.