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This study was aimed to investigate the expression of miR-651-3p in breast cancer and its role in regulating the expression of immunosuppressive factors IL-10 and TGFβ1 as well as the apoptosis level of breast cancer cells.qRT-PCR was used to detect the expression level of miR-651-3p in MCF-7 cells.miR-651-3p-over-expressng/-silencing plasmids and SP2-silencing plasmid were used to transfect MCF-7 cells.The expression levels of SP2,IL-10 and TGFβ1 were detected by Western blot assays.The apoptosis level of MCF-7 cells was detected by flow cytometry.The binding between miR-651-3p and SP2 was verified by dual luciferase gene report,and the binding of SP2 to IL-10 and TGFβ1 promoter region was verified by ChIP assay.Our results showed that miR-651-3p was down-regulated in breast cancer cells(P<0.05).The over-expression of miR-651-3p and the knockdown of SP2 significantly down-regulated the expression levels of IL-10 and TGFβ1 and promoted the apoptosis of breast cancer cells,while silenced miR-651-3p had the opposite effect.miR-651-3p binds to the SP2-3'-UTR end and down-regulates its expression.Based on the silencing of miR-651-3p,the silencing of SP2 can significantly reduce the influence of silenced miR-651-3p on the expression of IL-10 and TGFβ1 and apoptosis level in breast cancer cells.SP2 promotes the expression levels of IL-10 and TGFβ1 by binding to their promoters,respectively.Taken together,down-regulated miR-651-3p in breast cancer inhibited the expression of SP2 by binding to its 3'-UTR region,reduced the up-regulation effect of SP2 on the expression of IL-10 and TGFβ1,thus inhibiting the expression of immunosuppressive factors IL-10 and TGFβ1 and promoting apoptosis in breast cancer cells.
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A novel strain,which could use 2-hydroxypyridine (2HP) as the sole source of carbon,nitrogen,and energy,was isolated from petroleum-contaminated soil at the Liaohe estuarine wetland.Strain 2PR was identified as Arthrobacter based on the morphology and 16S rRNA gene sequence.The optimum growth and degradation condition upon 2PR is 30℃ and pH 7.0,respectively.Under this condition,2HP degradation rate were 97.34％,94.95％,94.48％ and 89.21％ with 2,4,6 and 8 mg/ml initial concentration of 2HP at 42,96,120 and 156 h,respectively.Strain growth and 2HP degradation kinetics studies indicated that the strain followed Logisitic model,which could provide a theoretical and technical reference for the biodegradation of 2HP.The color of strain 2PR culture upon 2HP-MSN changed from colorless to blue,and then turned to brown.The blue pigment,which was observed at the culture of strain 2PR,was identified as 4,5,4',5'-tetrahydroxy-3,3'-diazadiphenoquinone-(2,2') by high performance liquid chromatography (HPLC) and high-performance liquid chromatography-mass spectrometry (LC-MS) analysis.The LC-MS signal with m/z =249.1 was observed in resting cells reaction sample with 2HP as the substrate.The degradation of 2HP might be achieved by a dioxygenase to produce 2,3,6-trihydroxypyridine,which could transformed to the blue pigment spontaneously,and then 2,3,6-trihydroxypyridine was converted with an pyridine-ring cleavage reaction.Among all the reported strains,strain 2PR has the strongest tolerance and the highest 2HP degradation efficiency at present.The strain has a promising application potential for 2HP waste treatment.
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Objective To investigate the effect of ellagic acid extracted from gallnut on multiple myeloma SP2 / 0 cell line and related mechanisms. Methods Multiple myeloma SP2 / 0 cell line was treated for 48 h with different concentrations of ellagic acid in vitro. Cell morphology,proliferation,apoptosis and cell cycle were analyzed with microscope,MTT experiment and flow cytometry,respectively. Tumor cell proliferation and apoptosis-related gene expression of COX-2 were detected by Western blotting. Results Cell cycle was arrested at the G1 phase 48 h after treatment with ellagic acid, the cell in G1 were (55. 21±3. 01)% , (64. 48 ± 0. 43)% , (75. 10 ± 2. 46)% , respectively, with significant difference as compared with control group [(34. 04±1. 74)% ,P<0. 01]. Cell suppression rate (21. 18±5. 92)% ,(44. 58±3. 43)% and (70. 15±2. 90)% ,respectively, in 20,40 and 60 μg·mL-1 ellagic acid treatment groups. Compared with the control group,the differences were significant (P<0. 01). Cell apoptosis rate was (9. 60 ± 0. 56)% , (19. 30 ± 1. 51)% and (35. 10 ± 5. 26)% , respectively, in 20,40 and 60 μg·mL-1 ellagic acid treatment groups,with significant differences as compared with the control group[(3. 23±0. 85)% ,P<0. 01]. With the increase of drug concentration,COX-2 expression was decreased. Conclusion Ellagic acid can inhibit myeloma SP2 / 0 cell proliferation and promote apoptosis.
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Aims: This study was carried out to further characterize fungal species that could degrade 3-chloropropionic acid (3CP) as sole source of carbon and energy. Methodology and Results: Both fungi were able to grow on 3CP after 10 days on solid minimal media. Based on sequencing of its segment of 18S rRNA these isolates were identified as Mucor sp. SP1 and Trichoderma sp. SP2. The isolated strains were not able to grow on media plates containing 10 mM of 2,2-dichloropropionate (2,2DCP) as sole source of carbon. 3CP degradation was observed in liquid minimal medium containing 10 mM 3CP after 18 days culture period. The chloride ion released was detected in both growth medium containing Mucor sp. SP1 and Trichoderma sp. SP2. At least 80% of 10 mM 3CP was utilized in the growth medium. Conclusion, significance and impact of study: Dehalogenase enzyme that can degrade α-chloro-substituted haloalkanoic acids for example 2,2DCP is well studied up to protein crystallization. Very few reports on the degradation of β-chloro-substituted haloalkanoic acids such as 3CP and none from fungi. This study is considered important because it can be compared to that of well-documented α-chloro-substituted haloalkanoic acids degradation. This is the first study to indicate fungal growth on 3CP as sole carbon and energy sources.
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The gene expression of both the mouse plasmocytoma (SP2/0) and the hybrid cells crossed with rat nucleated erythroblasts were detected by in situ hybridization technique using the probes of mouse ?-globin gene and 7 oncogenes (v-Ki-ras, v-H-ras, v-sis, erb-B, v-abl, v-fos, c-myc). After plasmic amplification, DNA was isolated by alkali lysis, purified and recovered, the DNA containing gene fragments were labelled with ~(32)P to become high activity ~(32)P-cDNA probes through nick translation, and the labelled probes were used to detect the gene transcripts in cellular level. The results indicated that: (1) no mouse ?-globin gene transcripts could be detected in the cytoplasm of SP2/0 cells, as well as in hybrid cells within 72 hours after cell fusion, but transcript signals could be observed in hybrid cells from 4th to 26th passages. (2) Active expression of multioncogenes in SP2/0 cells was demonstrated, all the 7 oneogenes tested, except v-sis, were expressed more strongly. On the other hand, the expression of oncogenes in hybrid cells was found to be dramatically decreased, among them, the oncogenes of c-myc and Ki-ras been suppressed completely. After long term of passages (26th subcultures), the expression of c-myc and Ki-ras was still lower than that of SP2/o ceils although in some cases other oncogenes increased in their expression levels. These results confirmed that the multistep carcinogenesis involved multi-oncogenes expression and that the decancerization of tumor cells may be due to the suppression of multi-oncogenes activity as well as to activate the expression of differentiation genes.
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The present study reported the observations with light and electronic micros copy on hybrid cells crossed between rat intermediate or late erythroblasts and mouse SP2/0 plasmocytoma cells. In a short period after fusion, the cell size and the ratio of nuclear heterochromatin in hybrid cells appeared to be increased, but the number of nucleoli, as well as the number of microvilli, finger-like processes, and nuclear cytoplasmic ratio were decreased. Swelling mitochondria, pycnotic nuclei and/or process of enucleation also could be seen in some hybrid cells. The subcul tured hybrid cells were characterized with less microvilli and cellular surface membrane processes, and showing marked changes in nuclear size, as well as the appearance of cytoplasmic vesicles and dense granules in some cells. The observations mentioned above provide morphological and ultrastructural evidences for the regulation of malignant phenotype of hybrid cell model we reported previously. The possible relationship between the deeancerization and the morphological changes of hybrid cell nucleus, cytoplasm and cell surface were briefly discussed.