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OBJECTIVE To investigate the inhibitory effects of ursolic acid on interleukin-6 （IL-6）-mediated invasion and migration of breast cancer MDA-MB-231 cells （hereinafter referred to as “231 cells”）. METHODS The effects of 20， 40， 80， 160 and 320 µmol/L ursolic acid on the proliferation rate of 231 cells were measured by CCK-8 method. The breast cancer 231 cells were divided into control group， model group and administration group. The migration and invasion abilities of cells were detected by scratch assay and Transwell assay. Real-time quantitative polymerase chain reaction （q-PCR） assay and Western blot assay were used to detect the mRNA and protein expressions of epithelial-mesenchymal transition-related makers such as E cadherin （E-cad）， matrix metalloprotein 2 （MMP2）， MMP9， vimentin （Vim）， CD44 molecule （CD44） and aldehyde dehydrogenase 1 family member A1 （ALDH1A1）. The phosphorylation levels of JAK2 and STAT3 in the Janus kinase 2/signal transducer and activator of transcription 3 （JAK2/STAT3） signaling pathway （in terms of p-JAK2/JAK2 ratio and p-STAT3/STAT3 ratio） were detected by Western blot assay. RESULTS A low concentration of ursolic acid of 20 µmol/L （no significant inhibitory effect on cell proliferation ability） was selected as the subsequent administration concentration. Compared with the control group， the migration and invasion abilities of cells in the model group were significantly enhanced （P＜0.05）； compared with the model group， the migration and invasion abilities of cells in the administered group were significantly reduced （P＜0.05）. Compared with the control group， the relative mRNA and protein expressions of epithelial-mesenchymal transition-related markers MMP9， MMP2， Vim， ALDH1A1 and CD44 were all elevated to different extents， and the mRNA and protein expressions of E-cad were all decreased to different extents in the model group cells， and part of the differences had statistical significance （P＜0.05）， the p-JAK2/JAK2 ratio and p-STAT3/STAT3 ratio were significantly increased in the model group （P＜0.05）； compared with the model group， the expressions of the above indicators were reversed to some extent in the administration group. CONCLUSIONS Ursolic acid blocks the activation of JAK2/STAT3 signaling pathwby the inflammatory factor IL-6， which ultimately interrupts the invasion and metastasis of breast cancer cells.
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Myeloid differentiation factor 88 (MyD88) plays an important role in tumorigenesis,development and malignant transformation,also participates in the microenvironment,proliferation,apoptosis,invasion,metastasis and tumor resistance.MyD88 may serve as a new and meaningful therapeutic target,which can promote the growth and development of tumors by regulating multiple signaling pathways and enhance the drug resistance of tumor cells.
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Objective To investigate the effect of ursolic acid (UA) on the colorectal tumor and microenvironment in mice,and to provide a theoretical basis for the clinical application of UA.Methods The models of subcutaneous transplanted tumor of mouse CT26 cells was established.The models were divided into four groups:control group,tumor bearing group,tumor beating dimethyl sulfoxide (DMSO) group and tumor beating UA group.Tbe serum levels of interleukin-6 (IL-6) were detected by enzyme linked immunosorbent assay (ELISA).The number and percentage of myeloid-derived suppressor cell (MDSC) in the spleen of mice were analyzed by flow cytometry.The mRNA levels of IL-6 and signal transducer and activator of transcription 3 (STAT3) in tumor were examined by real-time quantitative polymerase chain reaction (RT-PCR).The protein levels of STAT3 and p-STAT3 in tumor were detected by Western blotting.Results The results showed that UA could significantly decrease the number of spleen MDSC.The accounts of spleen MDSC of tumor bearing UA group (249.60 ± 17.12) was lower than that of tumor beating DMSO group (366.40 ± 34.08),and the difference was statistically significant (P =0.021).The serum level of IL-6 in tumor bearing UA group [(46.40 ± 17.05) pg/ml] was decreased than that in tumor bearing DMSO group [(94.27 ±21.51) pg/ml],and the difference was statistically significant (P =0.012).The expression levels of IL-6 and STAT3 mRNA in tumor tissues of tumor bearing UA group (0.12 ±0.01,0.21 ±0.08) were lower than those of tumor bearing DMSO group (0.69 ± 0.14,0.79 ± 0.06),and the differences were statistically significant (P =0.008;P =0.003).The protein expression levels of STAT3 and p-STAT3 in tumor tissues of tumor bearing UA group (0.81 ±0.02,0.28 ±0.04) were lower than those of tumor bearing DMS0 group (0.98 ±0.02,0.91 ±0.22),and the differences were statistically significant (P =0.027;P =0.029).Conclusion UA may inhibit the activation of STAT3 signaling pathway and the amplification of MDSC in microenvironment by reducing IL-6,thus to enhance the function of immune-killing tumor cells to regulate tumor immune microenvironment and inhibit the immune escape of mouse colorectal cancer cells.
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Objective To prepare a peptide monoclonal antibody (McAb)against human papillomavirus 18E6 separately,and identify its specificity and pathogenicity.Metheds The advantage epitope peptide was designed and synthesized by ABCpred and Bcepred,and then used to immunize BALB/c mice after coupling with bovine serum albumin (BSA).And the McAb was prepared by hybridoma technique.HPV18E6 gene was amplified from cervical swab specimen containing HPV18 and insert-ed into expression vector pET-28a.The constructed recombinant plasmid was transformed to E.coli BL21(DE3)for expres-sion under induction of isopropyl thio-β-D-galactoside.The expressed protein was used to identified the McAb had been pre-pared.Results The hybridoma cell lines could constantly produce MAbs against HPV18E6 peptides.Sequencing proved that recombinant plasmid pET-28a-HPV18E6 was constructed correctly.Western blotting showed that the anti-HPV18E6 pep-tides antibody could specifically recognize HPV18E6.Conclusion A monoclonal antibody against the advantage epitope pep-tide of human papillomavirus 18E6 prepared could specifically recognize HPV18E6 specifically.