Background:Recent studies have shown that long non-coding RNAs(lncRNAs)play important roles in carcinogenesis and cancer biology and the related context has attracted more and more attentions. PVT1,which encodes a lncRNA,is reported to be up-regulated and exhibit pro-oncogenic activity in a wide variety of human cancers. Aims:To investigate the expression of PVT1 in human pancreatic cancer cells and its effect on proliferation and apoptosis of HPAF-Ⅱ cells. Methods:One target siRNA against PVT1 was synthesized and transfected into HPAF-Ⅱ cells by using lipofactamine technique. PVT1 mRNA expression was detected by real-time PCR;capability of cell proliferation was examined by MTS and colony formation assays;cell cycle progression and apoptosis were measured by flow cytometry;and Western blotting was performed to determine the expressions of apoptosis-related proteins and proto-oncogene protein c-Myc. Results:The mRNA expression of PVT1 in several human pancreatic cancer cell lines,especially HPAF-Ⅱ cells was significantly higher than that in H6c7,a human immortalization normal pancreatic ductal epithelial cell line. Compared with HPAF-Ⅱ cells transfected with negative control siRNA or without transfection,silencing of PVT1 by siRNA-PVT1 resulted in remarkable reduction in cell proliferation,cell cycle G1 phase arrest,and notable apoptosis;meanwhile,the expressions of apoptosis-related proteins(cleaved-caspase-3 and cleaved-PARP)were up-regulated,the ratio for Bcl-2 / Bax was decreased,and the expression of c-Myc protein was down-regulated. Conclusions:LncRNA-PVT1 is highly expressed in human pancreatic cancer cell line HPAF-Ⅱ. It may affect the proliferation and apoptosis of HPAF-Ⅱ cells partially through regulating c-Myc expression.

Objective To investigate the effects on self-renewal of pancreatic cancer stem cells by inhibiting hedgehog signaling pathway through cyclopamine. Methods PANC1 stem cells, PANC1 adherent cells and immortalized pancreatic ductal epithelial H6C7 cells were treated with 0.5, 1, 2, 5, 10 mol/L of cyclopamine for 24, 48, 72 h. The expression of Smo mRNA and Gli1 mRNA were detected by real-time PCR.Cell growth viability was measured by CCK 8. Cell cycle and apoptosis were determined by flow cytometry.Results Seventy-two hours after cyclopamine treatment, the Smo mRNA expressions of PANC1 stem cells,PANC1 adherent cells and H6C7 cells were 1,0.83 and 2.61; the expressions of Gli mRNA were 57.27,26.35,1; the inhibitory rates were ( 37.85 ± 13.69 ) %, ( 8.53 ± 4.43 ) %, (43.55 ± 28.98 ) %. Compared with PANC1, the expressions of Smo mRNA, Gli1 mRNA and the inhibitory rate of PANC1 stem cells significantly increased ( P ＜ 0.05 ). The proportion of G1 stage of PANC1 stem cells significantly decreased from (67.41 ±6.35)% to (36.53 ±6.03)% (P ＜0.05), and the apoptosis decreased from (10.95 ±5.68) % to ( 5.73 ± 1.42 ) % ( P ＞ 0.05 ). The proportion of G1 stage of PANC1 cells significantly decreased from ( 67.64 ± 6.88 ) % to ( 53.13 ± 1.10 ) % ( P ＜ 0.05 ); the apoptosis decreased from ( 12.08 ±4.12)% to (5.66 ± 1.33)% (P ＞0.05). While both the proportion of G1 stage and apoptosis of H6C7 cells was not significantly different. Conclusions Cyclopamine can inhibit the proliferation of PANC1 stem cells via blocking hedgehog signal pathway, and the mechanism may not be associated with cell apoptosis.

Objective To establish the method of suspension culture for stem cells from human pancreatic cancer cell line PANC1.Methods PANC1 cells were cultured in serum-free medium under floating-culture system.Tumor cell spheres were observed by optical microscope.Expression of CD133 and cell cycle were detected by flow cytometry.Cancer stem cells were induced to differentiate with 10%FBS,and expression of CK18,was evaluated with immunofluorescence microscope.Spheres cells were injected into the subcutaneous space of NOD/SCID rat and tumor formation was monitored weekly.Results PANC1 cells could form the stem cells spheres,and the rate of sphere formation was stable between 4%0 and 5%0 after 20 passages in vitro.The expression of CD133(5.91±0.7%)and proportion of G0/G1 phase cell(80.99±2.60%)was significantly increased in spheres cells compared with parental PANC1 cells(1.44±0.52%and 69.01±5.03%),and the difference was statistically significant(P＜0.05).When these spheres cells were cultured in media with serum,these cells gradually returned to the status of parental cells and expressed CK18,2×103 sphere cells injection could initiate tumor fornmtion in NOD/SCID rat.Conclusions Tumor spheres stem cellscould be generated under serum-free floating-culture system.The sphere cells possessed the capacities of self renew,difierentiation,and tumorigenic potential.

Objective: To investigate the characteristic and prognostic significance of leukemia stem cells associated antigens expressions including CD34, CD38, CD123, CD96 and TIM-3 in t (8;21) AML. Methods: Bone marrow samples of 47 t (8;21) AML patients were collected at diagnosis from October 2015 to April 2018 in Peking University Peoples’ Hospital, then flow cytometry method was performed to detect the expression frequencies of CD34, CD38, CD123, CD96 and TIM-3 to analyze the relationship between leukemia stem cells associated antigens expressions and relapse. Results: Of 47 t (8;21) AML patients tested, the median percentages of CD34⁺CD38^-, CD34⁺ CD38^-CD123⁺, CD34⁺CD38^- CD96⁺ and CD34⁺ CD38^- TIM-3⁺ cells among nucleated cells were 2.37%, 0.24%, 0.27% and 0.06%, respectively. All the frequencies of CD34⁺CD38^-, CD34⁺CD38^-CD123⁺, CD34⁺CD38^-CD96⁺ and CD34⁺ CD38^-TIM-3⁺ cells had no impact on the achievement of CR after the first course of induction. All higher frequencies of CD34⁺CD38^-, CD34⁺CD38^-CD123⁺, CD34⁺CD38^-CD96⁺ cells were related to higher 2-year CIR rate. Whereas, the frequency of CD34⁺ CD38^- TIM-3⁺ cells had no impact on CIR rate. Both high frequency of CD34⁺ CD38^- cells and the high level of minimal residual diseases (patients with <3-log reduction in the RUNX1-RUNX1T1 transcript level after the second consolidation therapy) were independent poor prognostic factors of CIR[P=0.025, HR=6.9 (95%CI 1.3-37.4) ; P=0.031, HR=11.1 (95%CI 1.2-99.2) ]. Conclusion Different leukemia stem cells associated antigens had distinct prognostic significance in t (8;21) AML. High frequencies of CD34⁺ CD38^-, CD34⁺ CD38^- CD123⁺ and CD34⁺CD38^-CD96⁺ cells at diagnosis predicted relapse in patients with t (8;21) AML.

【Objective】 To evaluate the safety and effectiveness of Perioperative Transfusion Trigger Score (POTTS) in guiding surgical patients blood transfusion intraoperatively and postoperatively. 【Methods】 A total of 900 patients(perioperative Hb 60~100 g/L) from December 2017 to March 2021 were collected, including 251 males and 649 females, with ASA grading Ⅰ~Ⅳ, and randomly divided into experimental group and controls. In the experimental group, the allogeneic RBC transfusion trigger(Hb threshold) and transfusion units in anemia patients was determined by POTTS. While those in the controls were decided by physicians according to current guidelines concerning transfusion. The proportion and units of allogeneic RBC transfusion, the incidence of postoperative complications, the mortality of hospitalization and discharge for 4 weeks, Hb value, healing of surgical incision, ICU admission rate and ICU length of stay, length of hospitalization, etc were recorded. 【Results】 The proportion of allogeneic RBC transfusion in the experimental group(35.3%)were less than the controls(42.2%)(P<0.05). The units of allogeneic RBC transfused, incidence of postoperative complications and mortality in hospital and 4 weeks after surgery, healing of surgical incision (grade A/ grade B/ grade C), the Hb level 24 h after surgery and at discharge, the ICU admission rate and ICU length of stay, length of hospitalization were not significantly different between the groups. 【Conclusion】 The peri-operative allogeneic red blood cells transfusion guided by POTTS can reduce the proportion of allogeneic RBC transfusion, and is safe and effective.