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Objective To investigate the role and potential mechanism of high mobility group box-1 protein (HMGB1) on improving the chemosensitivity of gemcitabine-resistant pancreatic cancer PANC1 ceils.Methods Gemcitabine-resistant pancreatic cancer PANC1 (PANC1-GR) cell line was established by using increased gradient concentration of gemcitabine.The si-HMGB1-PANC1 and si-HMGB1-PANC1-GR cells were established by the transfection with HMGB1 siRNA using liposome.The 50％ inhibitory concentration (IC50) and Resistance index (RI) of gemcitabine in 4 PANC1 cell lines with or without HMGB1 siRNA transfection were determined and calculated by CCK-8 assay.Western blot assay was used to detect the protein expression of HMGB1 in PANC1 and PANC1-GR cells and the expression of autophagy marker protein Beclin1 in the 4 PANC1 cell lines.Flow cytometry assay was used to evaluate the apoptosis rate of 4 pancreatic caner cell lines.Results The gemcitabine-resistant pancreatic cancer cell line PANC1-GR was successfully established,which could grow stably and passage in media with 100 μmol/L gemcitabine.The IC50 of gemcitabine in PANC1,PANC1-GR,si-HMGB1-PANC1,and si-HMGB1-PANC1-GR cells lines were (4.7 ±0.4) μmoL/L,(166.5 ± 13.6) μmol/L,(3.2 ± 0.3) μmol/L,and (52.4 ± 8.4) μmol/L,respectively.The IC50 in PANC1-GR wassignificantly higher than that in PANC1,while the IC50 in the transfected cells was significantly lower than that in untransfected cells,and the differences were both statistically significant (bothP ＜ 0.01).The RI value of gemcitabine in transfected and untransfected PANC1-GR cells was 35.4 and 16.4.The relative protein levels of HMGB1 in PANC1 and PANC1-GR were 0.17 ± 0.08 and 0.38 ± 0.11.The expression of HMGB1 in PANC1-GR was obviously higher than that in PANC1,and the difference was statistically significant (P＜0.01).The relative protein levels of Beclin1 in PANC1,PANC1-GR,si-HMGB1-PANC1 and siHMGB1-PANC1-GR cells were 2.68 ± 0.23,3.28 ± 0.15,0.68 ± 0.23 and 0.78 ± 0.11,which in two transfected cells was greatly lower than those in untransfected cells.The apoptosis level was (34.58± 3.14)％,(79.56±3.58)％,(19.41± 1.53)％,and (34.57±2.94)％.The apoptosis level in the 2 transfected cell lines were significantly higher than those in the 2 untransfected cell lines,and the differences were both statistically significant (P ＜ 0.01).Conclusions The inhibition of HMGB1 could improve the chemosensitivity of gemcitabine in pancreatic cancer PANC1 cells,which might be mediated by the activation of autophagy.
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Objective To observe the regulatory role of long non-coding RNA HIF1A-AS1 on the autophagy of pancreatic cancer PANC1 cells induced by hypoxia.Methods The pancreatic cancer PANC1 cells were cultured in a three-gas incubator filled with hypoxic gas mixture (94% N2,5% CO2,1% O2) for 3, 6, 12, 24, 36 and 48 h.HIF1A-AS1 overexpression and low expression PANC1 cells were obtained by the infection of recombinant adenovirus carrying HIF1A-AS1 and the transfection of HIF1A-AS1 targeting siRNA by liposome, and routinely cultured PANC1 cells served as control.The expression of HIF1A-AS1 of PANC1 cells was detected by real-time quantitative PCR after being cultured in hypoxia-induced condition for 24 h.The apoptosis rate was detected by flow cytometry.The autophagy related proteins Beclin 1 were detected by western blot.Results The expression of HIF1A-AS1 in hypoxic cells was increased as the hypoxic time increased since 6 h and peaked at 36 h, which was significantly higher than that in control group (P<0.01).HIF1A-AS1 relative expression in HIF1A-AS1 overexpression and low expression PANC1 cells was 4.49±0.53 and 0.49±0.07, which were normalized to that of control group with the relative expression of 1.Control group had lower HIF1A-AS1 expression than HIF1A-AS1 overexpression PANC1 cells but higher HIF1A-AS1 in HIF1A-AS1 low expression PANC1 cells, and the differences were statistically significant (P<0.01).The cell apoptosis rate of control, HIF1A-AS1 overexpression and low expression PANC1 cells was (8.27±1.28)%, (6.56±1.49)% and (19.9±2.34)% after 24 h hypoxic culture.Control group had higher HIF1A-AS1 expression than HIF1A-AS1 overexpression PANC1 cells but lower HIF1A-AS1 in HIF1A-AS1 low expression PANC1 cells, and the differences were statistically significant (P<0.01).The expression of Beclin 1 protein was protein 1.05±0.11, 1.29±0.19 and 0.38±0.18, respectively.Control group had lower Beclin 1 expression than HIF1A-AS1 overexpression PANC1 cells but higher Beclin 1 in HIF1A-AS1 low expression PANC1 cells, and the differences were statistically significant (P<0.01).Conclusions HIF1A-AS1 can promote autophagy of pancreatic cancer PANC1 cells induced by hypoxia and participate in the pathogenesis and metastasis of pancreatic cancer.
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Twenty six patients with fracture of tibial plateau was under arthroscopy assisted reduction, the joint surface of bone graft, and USES the steel plate fixation treatment. Average surgery time was 65 min (70-120 min), average fracture healing time was 15 weeks (12-17 weeks), joint surface anatomical reattachment rate was 92.9%. Using break knee function criteria evaluation of curative effect: 18 cases great 6 cases wed, 2 cases ok, fine rate was 92.3%. No infection, deep venous thrombosis and small leg fascia chamber syndrome and other complications. Conclusion is that treatment of tibial plateau fractures under arthroscope has advantages of small trauma, check intuitively and reset accurately, functional recovery of patients are satisfied, the treatment has certain clinical application value.
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Adult , Female , Humans , Male , Middle Aged , Young Adult , Arthroscopy , Minimally Invasive Surgical Procedures , Tibial Fractures , General Surgery , Treatment OutcomeABSTRACT
Objective To investigate the expression of miR-101 in pancreatic cancer and the effect of down-regulation miR-101 on proliferation of pancreatic cancer cell line ASPC1. Methods Real-time PCR was used to determine the expression of miR-101 in pancreatic cancer, adjacent tissues and pancreatic cancer cell line ASPC-1. The miR-101 over-expression vector (peGFPc1-miR-101) was constructed and was transfected into ASPC-1 cell. Transfection efficiency was measured by fluorescence microscope. The expression of miR101in the transfected cells was detected by real-time PCR. Cell viability analysis was performed by MTT. The targeted genes of miR-101 in pancreatic cancer were scanned by the online targeted gene prediction software (target Scan). Results The expression of miR-101 was in pancreatic cancer tissues, adjacent tissues and ASPC-1 cell line, respectively. The expressions in pancreatic cancer tissues and ASPC-1 cells were significantly lower than that in adjacent tissues ( P ＜ 0.01 ). The expression of miR 101 in transfected cells increased to 19.8 folds as much as that in the control group (P ＜0.01 ). Proliferation rate of transfected cells was significantly decreased, which was only 26% of primary cells ( P ＜ 0.01 ). EZH2 was the potential targeted gene of miR-101 in pancreatic cancer. Conclusions miR-101 was weakly expressed and it may affect the proliferation of pancreatic cancer cell by inhibiting the EZH2 expression.