[Effects of tumor protein translation control antisense RNA1 on radiosensitivity, proliferation, migration and invasion of hepatocellular carcinoma cells by targeting miR-30c-5p].

Objective: To investigate the effects of tumor protein translation control antisense RNA1 (TPT1-AS1) on the radiosensitivity, cell proliferation, migration and invasion of hepatocellular carcinoma cells by targeting microRNA-30c-5p (miR-30c-5p). Methods: Thirty-four cases of liver cancer tissues and adjacent normal tissues were derived from liver cancer patients who were admitted to Shanxi Provincial People's Hospital from March 2016 to March 2018. Liver cancer HepG2 cell was transfected with negative control siRNA (si-NC group), si-TPT1-AS1 (si-TPT1-AS1 group), pcDNA3.1 (pcDNA3.1 group), pcDNA3.1-TPT1-AS1 (pcDNA3.1-TPT1-AS1 group), si-TPT1-AS1 and anti-miR-NC (si-TPT1-AS1+ anti-miR-NC group), si-TPT1-AS1 and anti-miR-30c-5p (si-TPT1-AS1+ anti-miR-30c-5p group), respectively. Real-time quantitative reverse transcription polymerase chain reaction (qPCR) was used to detect the transcription levels of TPT1-AS1 and miR-30c-5p in normal tissues adjacent to cancer and liver cancer tissues, the clone formation test was used to test the radiosensitivity of HepG2 cells, and the Methyl Thiazolyl Tetrazolium (MTT) test was used to test the proliferation of HepG2 cells. Cell cycle distribution was detected by flow cytometry, Transwell array was used to detect the migration and invasion ability of HepG2 cells, dual luciferase reporter array was used to verify the targeting relationship of TPT1-AS1 and miR-30c-5p, western blot was used to detect the expressions of proliferation, migration and invasion-related proteins. Results: The expression levels of TPT1-AS1 and miR-30c-5p in liver cancer tissues were 0.84±0.08 and 0.13±0.01, statistically different from 0.31±0.03 and 0.50±0.05 in normal tissues adjacent to cancer (P<0.05). When the cells were treated with 2, 4, 6, 8 Gy irradiation, the cell survival scores of the si-TPT1-AS1 group were 0.280±0.040, 0.069±0.011, 0.020±0.003 and 0.005±0.001, respectively, lower than 0.648±0.070, 0.348±0.080, 0.130±0.020 and 0.060±0.009 of the si-NC group (P<0.05), the radiosensitization ratio of the si-TPT1-AS1 group was 1.672. The number of cell migration and invasion in the si-TPT1-AS1 group were (50.00±4.36) and (44.00±4.03), respectively, which were lower than (109.00±8.68) and (94.00±7.49) in the si-NC group (P<0.05), the cell absorbance (A) values at 24, 48 and 72 hours were 0.28±0.03, 0.43±0.04 and 0.68±0.07, respectively, lower than 0.46±0.04, 0.87±0.08 and 1.35±0.13 of the si-NC group (P<0.05), the protein expression levels of Cyclin D1, p21, E-cadherin and MMP-2 were 0.25±0.02, 0.65±0.06, 0.68±0.07 and 0.27±0.03, respectively, statistically different from 0.88±0.08, 0.17±0.02, 0.14±0.01 and 0.89±0.09 of si-NC group (P<0.05), the proportions of S phase and G(2) phase in the si-TPT1-AS1 group were (17.82±1.03)% and (34.15±2.29)%, respectively, significantly different from (35.14±2.61)% and (16.84±1.21)% in the si-NC group (P<0.05). The luciferase activity of cells in the WT-TPT1-AS1+ miR-30c-5p group was 0.26±0.02, lower than 0.92±0.09 in the WT-TPT1-AS1+ miR-NC group (P<0.05). The cell survival scores in the si-TPT1-AS1+ anti-miR-30c-5p group were 0.450±0.081, 0.200+ 0.045, 0.070±0.010, 0.026±0.004 after treatment with 2, 4, 6, 8 Gy irradiation, higher than 0.285±0.043, 0.075±0.014, 0.028±0.004, 0.006±0.001 of si-TPT1-AS1+ anti-miR-NC group (P<0.05). The radiosensitization ratio of the si-TPT1-AS1+ anti-miR-30c-5p group was 0.694. The number of migration and invasion in the si-TPT1-AS1+ anti-miR-30c-5p group were 79.00±6.65 and 68.00±6.33, higher than (52.00±4.41) and (46.00±4.06) of si-TPT1-AS1+ anti-miR-NC Group (P<0.05), the A values at 24, 48 and 72 hours were 0.37±0.03, 0.64±0.06 and 0.96±0.09, respectively, higher than 0.26±0.03, 0.41±0.04, and 0.65±0.06 of si-TPT1-AS1+ anti-miR-NC group (P<0.05), the expression levels of Cyclin D1, p21, E-cadherin and MMP-2 protein were 0.57±0.06, 0.43±0.04, 0.43±0.04 and 0.64±0.06, statistically different from 0.24±0.02, 0.66±0.06, 0.65±0.06 and 0.28±0.03 of the si-TPT1-AS1+ anti-miR-NC group (P<0.05). Conclusions: The expression of TPT1-AS1 up-regulates in the liver cancer tissues. TPT1-AS1 may down-regulate miR-30c-5p expression, reduce the radiosensitivity of liver cancer cells, and promote the proliferation, migration and invasion of liver cancer cells.

[Effect of lncRNA HOTAIR on the radiosensitivity of HCCLM3 cells].

Objective: To investigate the effect of down-regulating long non-coding RNA (lncRNA) and HOX transcript antisense RNA (HOTAIR) targeting miR-761 on the radiosensitivity of HCCLM3. Methods: The expression of HOTAIR in liver cancer cells HuH-7, SNU-449, HCCLM3 and normal liver cells L-02 were measured by real-time quantitative PCR. HCCLM3 cells were divided into control, Sh-NC (transfected shRNA negative control), Sh-HOTAIR (transfected HOTAIR shRNA), RAD+Sh-NC (transfected shRNA negative control and irradiated with 8Gy dose), and RAD+Sh-HOTAIR (HOTAIR shRNA was transfected and irradiated with 8Gy dose) group. Apoptosis was detected by flow cytometry, Bcl-2 Associated X Protein (Bx-2), cleaved cysteine-containing Cleaved cysteinyl aspartate specific proteinase 3 (C-Caspase-3) protein expression. Sh-NC, Sh-HOTAIR cells were irradiated with 0, 2, 4, 6, 8 Gy, and plate-clone experiments were used to determine radiosensitivity. Bioinformatics software predicted that miR-761 might be a target gene of HOTAIR, and the luciferase reporter system identified the targeting relationship. The miR-761 inhibitor, HOTAIR shRNA and inhibitor negative control, and HOTAIR shRNA were co-transfected into HCCLM3 cells, respectively. Cell apoptosis and Bax and C-cysteine-containing aspartate proteins were also measured using the above method, as well as the hydrolase-3 protein expression and cell survival fraction. Results: The expression levels of HOTAIR in liver cancer cells HuH-7, SNU-449, and HCCLM3 were higher than those in normal liver cells L-02 (1.85±0.12, 2.27±0.23, 2.68±0.15 vs 1.00±0.09, P<0.05). Compared with Sh-NC, the apoptosis rate of Sh-HOTAIR, RAD+Sh-NC cells and Bax, C-Caspase-3 protein levels are higher [Apoptotic rate: (13.47±1.32)%, (12.84±1.19)% vs (2.98±0.27)%; Bax protein: 0.74±0.08, 0.72±0.06 vs 0.42±0.06; C-Caspase-3 protein: 0.56±0.06, 0.54±0.08 vs 0.25±0.04, all P<0.05]. Compared with Sh-HOTAIR and RAD+Sh-NC, RAD+Sh-HOTAIR cell apoptosis rate and Bax, C-Caspase-3 protein levels are higher [apoptosis rate:(22.57±2.36)% vs (13.47±1.32)%, (12.84±1.19)%, Bax protein: 0.99±0.11 vs 0.74±0.08, 0.72±0.06, C-Caspase-3 protein: 1.03±0.12 vs 0.56±0.06, 0.54±0.08,all P<0.05]. Compared with Sh-NC, Sh-HOTAIR cells had lower survival scores and higher radiosensitivity (P<0.05). HOTAIR targets negative regulation of miR-761 expression. Compared with cells co-transfected with inhibitor negative control and HOTAIR shRNA, cells co-transfected with miR-761 inhibitor and HOTAIR shRNA had lower apoptosis rate after radiation treatment [(10.24±1.32)% vs (21.84±2.01))%], Bax (0.50±0.06 vs 1.01±0.10) and C-Caspase-3 protein (0.56±0.07 vs 1.05±0.14) had lower expression and higher cell survival scores (all P<0.05). Conclusion: Down-regulating lncRNA HOTAIR targets miR-761 to increase the radiosensitivity of HCCLM3.

[Analysis of the entrance and suture method of umbilical incision in gynecological laparoscopy].

Objective: To explore the entrance and suture method of umbilical incision in gynecological laparoscopy. Methods: A total of 204 cases of gynecologic laparoscopy in our hospital from 2013 to 2016 were reviewed respectively. All the cases used two kinds of approach of umbilical incision: intra-umbilical incision and peri-umbilical incision (longitudinal/transverse oblique/arc incision according to the bellybutton natural skin folds) and two methods of suture: the suture of "U" and the suture of the whole subcutaneous tissue. Two groups were randomly assigned based on the entrance and suture method with each group 102 cases. The peri-operative outcomes were compared, including intra-operative and postoperative bleeding, postoperative incision fat liquefaction and infection, incision pain, incision appearance satisfaction and incision healing satisfaction. Results: The difference was statistically significant in the intra-operative and postoperative bleeding between two groups of intra-umbilical incision and peri-umbilical incision (P<0.05). There was statistically significant difference in postoperative incision fat liquefaction, infection and incision pain between two groups of the suture of "U" and the suture of the whole subcutaneous tissue (P<0.05). The incision appearance satisfaction showed no difference (P>0.05), but the difference was statistically significant (P<0.05) in the incision healing satisfaction between two groups. Conclusion: The peri-umbilical incision (longitudinal/transverse oblique/arc incision according to the bellybutton natural skin folds) and suture of the whole subcutaneous tissue can be the feasible modified methods with high practicability and security, good cosmetic result. It should be promoted in gynecologic laparoscopy.

[The expressions and the roles of SDF-1/CXCR-4 and SDF-1/CXCR-4 in human endometriosis].

Objective: To investigate the expressions and significances of SDF-1 and its receptor CXCR-4 in endometriosis. Methods: 50 hysteromyoma patients treated at Beijing Tongren Hospital, Capital Medical University between 1(st) January 2016 and 31(st) December 2017 were divided into control group, that is, non-endometriosis group, while another 50 endometriosis patients were divided into experimental group.The endometrial tissues, endometriosis lesions, and peritoneal fluid samples of hysteromyoma patients were collected by operation.RT-PCR, ELISA and immunohistochemistry were adopted to detect the expressions of SDF-1 and CXCR-4 in the two groups.Cell count was used to analyze the roles of SDF-1 and CXCR4 in the mitosis and proliferation of endometrial stromal cells. Results: The mean value of SDF-1 and CXCR-4 expressions in ascites or peritoneal fluid of endometriosis patients were (2.56±0.33) mg/L and (4.47±0.32) mg/L, respectively. The mean concentrations in ascites or peritoneal fluid in hysteromyoma patients were (1.39±0.36) mg/L and (3.16±0.32) mg/L, respectively.The expressions of SDF-1 and CXCR-4 in ascites or peritoneal fluid of endometriosis patients were both significantly higher than those of patients in non-endometriosis group (P<0.05). SDF-1 and CXCR-4 were expressed in both endometriosis lesions and the glandular epithelial cells and mesenchymal cells of normal endometrial tissue.Positive staining sites were located in the cytoplasm.A value was used to calculate and analyze the expression of immune staining.The mean A value of SDF-1 and CXCR-4 in endometriosis group were 0.21±0.13 and 0.21±0.13, respectively, and the mean A value in normal endometrial tissues were 0.15±0.13 and 0.14±0.13, respectively.The expressions of these two in endometrial tissues were significantly higher than that in normal endometrial tissues, and the differences were statistically significant (P<0.05). The expression of CXCR-4 mRNA was abundant in the mesenchymal cells of endometriosis cultured in vitro.SDF-1 promoted the mitosis and proliferation of endometrial stromal cells cultured in vitro in a dose-dependent manner.The neutralizing antibody against CXCR-4 was obviously inhibited. Conclusion: The high expressions of SDF-1 and CXCR-4 in endometriosis as well as SDF-1 through its specific receptor CXCR-4 promoted the mitosis and proliferation of endometrial stromal cells, suggesting that SDF-1 and CXCR-4 played important roles in the pathogenesis of endometriosis.

New iridoids from Pedicularis artselaeri.

Three new iridoids, named artselaenin I, II and III, were isolated from the whole plants of Pedicularis artselaeri, along with 11 known compounds, 8-epiloganic acid, 7-deoxy-8-epiloganic acid, plantarenaloside, mussaenoside, lariciresinol-4-O-beta-D-glucoside, lariciresinol-4'-O-beta-D-glucoside, alaschaniosideA, cirtusinA, 2-(p-hydroxyphenyl)-ethanol 1-O-beta-D-glucopyranoside, 3-methoxy-4-primeverosylacetophenone and adenine. Their structures were identified mainly by spectral evidence.

[Study on the mass spectrometry of natural products. XI. The MIKES of fast atom bombardment mass spectra of sodium adduct ions of phenylpropanoid glycosides].

The fast atom bombardment (FAB) mass spectra and mass-selected ion kinetic energy spectra [MIKES] of sodium adduct ions [M + Na]+ of six phenylpropanoid glycosides have been studied. An intense sodium adduct appeared in their FAB mass spectra, which was more abundant than the corresponding protonated molecular ion [M + H]+ and may give more definite information about molecular weight of the sample. The MIKES of these [M + Na]+ ions provided a convenient way for sugar sequence analysis of such glycosides.