MicroRNA-375 regulates proliferation and apoptosis of glioma cancer cells by inhibiting CTGF-EGFR signaling pathway.

AIM: To evaluate the correlation between miRNA-375 and cell proliferation and apoptosis in glioma cancer cell. METHODS: Collecting 30 cases of glioma cancer patients and 30 cases of cerebral infarction patients. The miRNA-375 and CTGF protein expressions were evaluated by ISH and IHC methods. In the cell experiment, the U87 cells were divided into 3 groups: NC group (the cells were treated with normal method); BL group (the cells were transfected with empty vector) and miRNA group (the cells were transfected with miRNA-375). The U87 cell proliferation and apoptosis rates and cell cycle of the different groups were measured by MTT and flow cytometry. The relative proteins (CTGF, EGFR, AKT, Erk and P21) expressions were measured by WB assay. RESULTS: The miRNA-375 and CTGF expressions of glioma cancer tissues were significantly different compared with those of no-cancer tissues (p < 0.05, respectively). In the cell experiments, the cell proliferation of miRNA group was significantly decreased compared with that of NC group (p < 0.05); the cell apoptosis and G1 phase rate of miRNA group was significantly decreased compared with NC group (p < 0.05, respectively). Depending on the WB assay, the CTGF, EGFR, AKT, Erk and P21 proteins expressions of miRNA group were significantly different compared with proteins expressions of NC group (p < 0.05, respectively). CONCLUSION: miRNA-375 over-expression suppresses glioma cancer cells development via CTGF-EGFR pathway (Fig. 3, Ref. 30).

Screening and identification of a tumor specific methylation phenotype in the colorectal laterally spreading tumor.

OBJECTIVE: We screened and identified the differential expression of the methylation phenotype in the whole genome of colorectal laterally spreading tumor (LSTs). MATERIALS AND METHODS: 3 tissue samples of colorectal polypoid adenomas (PAs), 3 tissue samples of LSTs and 3 tissue samples of colon cancer were analyzed with a high-density gene chip, and about 450,000 methylation sites were detected covering approximately 95% of the CpG islands. The Delta Data screening was taken through a cluster analysis of methylation phenotype differential expression. 50 tissue samples each of PAs patients, LSTs patients, and colorectal cancer patients were selected. Methylation-specific PCR (MSP) was used to detect RASSF1A and WIF-1 methylation levels. He RT-PCR method was used to detect the relative mRNA expression levels for methylation expression identification. RESULTS: The degree of LST methylation was higher than that of PAs, and 1234 genes were found to have a lower expression when compared to colorectal cancer samples. 764 genes had a higher expression when compared to colorectal cancer, and 559 genes lower expression when compared to PAs. The average methylation level of LSTs was higher than that of PAs, and lower than that of colorectal cancer. The chromosomal location was taken on these 1234 genes, which were higher than that of PAs, and lower than that of colorectal cancer; 518 genes were located on chromosome No. 2 (41.98%), 236 on No. 5 (19.12%), 357 on No. 8 (28.93%), and 123 on No. 10 (9.97%). According to clustering analysis, DNA differentially methylated sites were mainly on genes of cell adhesion molecules regulation, signaling pathways, energy transduction, cell cycle and apoptosis. The positive rate of RASSF1A and WIF-1 methylation in the tissues of LSTs patients were higher than that of PAs, and lower than that of colorectal cancer; differences were statistically significant (p<0.05). The relative expression levels of RASSF1A and WIF-1mRNA in the tissues of LSTs patients were lower than that of PAs, higher than that of colorectal cancer, and the difference was statistically significant (p<0.05). CONCLUSIONS: The administration of high-density gene chip technology has a good application value to screen the differential expression of LSTs gene methylation phenotype. Results are consistent with the identification results.

[miR-202 contributes to sensitizing MM cells to drug significantly via activing JNK/SAPK signaling pathway].

Objective: To explore the role of miR-202 in multiple myeloma (MM) cells, and study the regulation of miR-202 on drug sensitivity of MM cells. Methods: miR-202 and BAFF mRNA levels were detected by real-time PCR. U266 cells were transfected with miR-202-mimics, miR-202-inhibitor, siBAFF and their negative controls. After above treatments, protein levels of Bcl-2 family and MAPK signaling pathway were detected by Western blot analysis, and the proliferation and apoptosis ability of MM cells were examined by WST-1, Annexin V-FLUOS assay, respectively. Results: The results showed that the expression of miR-202 in CD138+ MM cells (0.304±0.354) and U266 cells (0.052± 0.009) were lower than in normal controls (3.550 ± 1.126) (P<0.001, P=0.009), whereas BAFF mRNA levels (5.700 ± 0.734, 9.576 ± 2.887) were higher than in normal controls (1.819 ± 0.853) (P<0.001, P= 0.006). The proliferation ability of U266 cells transfected with miR-202 mimics was significantly inhibited than in control group [(56.04±0.021)% vs (18.89±0.32)%, P=0.002]. The result of Western blot showed that the expression of Bcl-2 decreased by about 24%, and the expression of Bax increased by about 124% in cells transfected with miR-202 mimics. The apoptosis rate in cells transfected with miR-202 mimics was significantly more than in control group [(49.60±4.89)% vs (26.20±1.28)%, P=0.029]. The apoptosis rate in miR-202 mimics combined with Bort group (51.23±5.41)% was higher as compared with Bort treatment alone (31.70±4.40)% or miR-202 mimics control combined with Bort group (27.94±4.04)%, (P=0.047, P= 0.028), whereas the apoptosis rate in miR-202 mimics combined with Thal or Dex had no significant difference compared with miR-202 mimics control [(11.66±1.91)% vs (10.63±1.74)%, P=0.700; (16.35± 1.32)% vs (17.43 ± 1.95)%, P=0.400]. The inhibitory rate of cell growth in miR-202 mimics combined with Bort group was higher as compared with Bort treatment alone [(36.93±5.98)% vs (18.18±4.10)%, P= 0.029]. The expressions of p-JNK protein decreased in U266 cells transfected with miR-202 mimics and treated with Bort. Conclusion: miR-202 mimics combined with Bort could inhibit proliferation and induce apoptosis of U266 cells through negative regulating target gene BAFF, which further inhibited the JNK/SAPK signaling pathway.

Survey of helminths in adult sheep in Heilongjiang Province, People's Republic of China.

The prevalence of helminths in adult sheep was investigated in Heilongjiang Province, People's Republic of China between January 1999 and September 2003. A total of 326 adult sheep representing local breeds (Xingjiang Fine Wool Sheep, Dongbei Fine Wool Sheep) as well as introduced breeds (Merino and Charollais) from representative geographical locations in Heilongjiang Province were slaughtered and examined for the presence of helminths. The worms were examined, counted and identified to species according to existing keys and descriptions. A total of 26 helminth species were found representing 2 phyla, 3 classes, 13 families and 20 genera. All sheep were infected by more than one helminth species. Oesophagostomum columbianum, Haemonchus contortus and Trichostrongylus colubriformis were the most common nematode species, and Paramphistomum cervi, Orientobilharzia turkestanica and Fasciola hepatica were the most common trematode species, whereas the infection of adult sheep with cestodes was uncommon. The results of the present investigation provide relevant "base-line" data for Heilongjiang Province, China, for assessing the effectiveness of future control strategies against helminth infections in sheep.