Objective To investigate the optimal cryopreservation liquid of the stable transfected cell line named hepatic carcino-ma HepG2 transfected by lentivirus .Methods Glycerol and DMSO as cryopreservation liguid ,The HepG2 transfected by lentivirus and HepG2 cells were both cryopreserved with four various proportion of cryopreservation liquid .After thawing ,the survival rate of the two cells were observed by inverted contrast microscope and the viability and proliferation were detected with MTT assay .Re-sults Using glycerol as cryoprotectant liquid ,the survival rate of the HepG2 cells transfected by lentivirus was obviously higher than other 3 groups when the proportion of the cryopreservation liquid(DMEM ∶FBS∶Glycerol)was 0∶9∶1(P=0 .001) ,where-as there was no significant difference among the groups of HepG2 cells with various proportion of cryopreservation liquid (P=0 .293) .Using DMSO as cryoprotectant liquid ,the survival rate of the cells transfected by lentivirus was 0% ,whereas the survival rate of the HepG2 cells was higher than 60% ,and there was no significant difference between groups of various proportion of cryo-preservation liquid(P=0 .487) .The MTT assay demonstrated that the higher the serum levels was ,the better proliferation capacity those cells had(P<0 .05) .Conclusion The optimal cryopreservation liquid for the hepatic carcinoma cell line HepG2 transfected by lentivirus is used glycerol as cryoprotectant solution ,and the proportion of cryopreservation solution was 90% FBS + 10%Glycerol .

Objective To research the expression of Sp3 andβ‐Catenin in HCC and study the assessable factors of them for prognosis in patients with hepatocellular carcinoma .Methods Western blot and RT‐PCR methods were used to detect the expres‐sion of Sp3 andβ‐Catenin in HCC and the liver tissue beside tumor among 49 cases .We analyzed the difference of these two indexes expressed in HCC and the liver tissue beside tumor .Then we detected the correlation between these two indexes and the character of clinic pathology ,and researched the correlation between Sp3 and the prognosis of HCC .Results The high expression rate of Sp3 in HCC was higher than that of liver tissue beside tumor(P<0 .05) according to Western blot and RT‐PCR ,the same toβ‐Catenin (P<0 .05) .Expression of Sp3 andβ‐Catenin were both related with size of tumor and degree of differentiation .Positive correlation existed between these two indexes according to Western blot method(r=0 .681 ,P=0 .000) and RT‐PCR method(r=0 .641 ,P=0 .000) .The prognosis of cases with high expression of Sp3 was poorer than the low expression cases(P<0 .05) .Conclusion Sp3 plays a promoter role in occurrence of HCC ,which is correlated with the grade malignancy of HCC .Sp3 might participated in occur‐rence and development of HCC via the Wnt pathway .

Objective To investigate the effect of deltaNp63(ΔNp63) silencing on the proliferation of pancreatic cancer PANC1 cells.Methods ΔNp63 mRNA level in 23 pairs of pancreatic cancer and adjacent tissue specimen was detected by real-time PCR, andΔNp63 protein in human normal pancreatic ductal cell line HPDE6-C7 and pancreatic cancer cell line PANC1, CFPAC1 and BXPC3 was detected by Western blot. PANC1 cells were transfectedΔNp63 specific siRNA (ΔNp63-siRNA ) and scramble siRNA ( Con-siRNA ) using liposome, and untransfected cells served as control.ΔNp63 mRNA and protein was detected by real-time PCR and Western blot to validate the silencing ofΔNp63 expression.MTT assay and BrdU method were used to detect the proliferation and DNA synthesis of transfected PANC1 cells.Results TheΔNp63 mRNA expression in pancreatic cancer tissues and matched adjacent normal tissues was 0.99 ± 0.07 and 0.70 ±0.07, respectively.ΔNp63 mRNA expression in pancreatic cancer tissue was significantly up-regulated compared with that in the normal tissue (P=0.0034).TheΔNp63 protein expression in HPDE6-C7, PANC1, CFPAC-1 and BxPC3 cells was 0.97 ±0.09,3.06 ±0.16,2.57 ±0.11 and 2.45 ±0.08, respectively.TheΔNp63 protein level in pancreatic cancer cells were higher than that in HPDE6-C7 cells (P<0.001).ΔNp63 mRNA level in control, Con-siRNA and ΔNp63-siRNA group was 0.97 ±0.07,0.97 ±0.07 and 0.28 ±0.03, respectively, andΔNp63 protein expression level was 0.97 ±0.06,1.00 ±0.10 and 0.26 ±0.03.The expression ofΔNp63 mRNA and protein inΔNp63-siRNA group were significantly down-regulated comparing with those in Con-siRNA group (P<0.01).Significant inhibition on cell proliferation was observed in ΔNp63-siRNA group, which was statistically different from that in control and Con-siRNA group.The A490 value (DNA synthesis) of control, Con-siRNA andΔNp63-siRNA group was 0.55 ±0.04, 0.56 ±0.01 and 0.55 ±0.00 at 24 h after transfection, and 0.84 ±0.05,0.87 ±0.07 and 0.71 ±0.05 at 48 h after transfection.The DNA synthesis inΔNp63-siRNA group was significantly down-regulated compared with that in control and Con-siRNA group (P<0.05).Conclusions Knockdown ofΔNp63 could greatly inhibit the proliferation and DNA synthesis of pancreatic cancer PANC1 cells.

Objective To investigate effect of taurine on respiration chain enzyme activity of mitochondria 24 hours after severe traumatic brain injury (TBI) in rats.Methods Fifty-six SD rats were divided into sham group,TBI group,taurine treatment group,and taurine prevention group according to the random number table,with 14 rats per group.Fluid percussion brain injury models were used.Via the caudal vein,normal saline was administered to animals in sham and traumatic brain injury groups immediately after injury,while taurine (200 mg/kg)was administered to animals in taurine treatment group after injury and in taurine prevention group 4 days before injury.Brains were harvested 24 hours postinjury for assays of HE staining and electron microscopy.Mitochondrial respiratory chain complex Ⅰ-Ⅴ activities were detected.Results TBI group presented swelling neurocytes,cell loss,karyopyknosis,shortened even vanished process,and inflammation cell infiltration at the edge of necrosis in HE staining.By contrast,morphological improvement was significant in taurine treatment group but only some neurons were intact in taurine prevention group.Swelling mitochondria and broken or vanished mitochondrial crests were seen in TBI group under the electron microscope.However,normal or minor swelling mitochondria was seen in taurine treatment group and cytoplasm slightly porous and absence of mitochondrial crests were seen in taurine prevention group.Activities of complex Ⅰ,Ⅱ and Ⅴ were significant lower in TBI group (32.52±2.41,4.68 ±0.15,2.49 ±0.73) compared to those in sham group (34.03 ±0.46,5.04 ±0.29,3.20±0.68) and in taurine treatment group (33.95±0.85,5.12-±0.23,3.53 ±0.48) (P＜0.05).And complex Ⅰ in taurine prevention group was significantly enhanced as well (34.44 ± 0.36,P ＜ 0.05).Conclusion Taurine may protect the brain tissues and mitochondrial structure from impairment in TBI rats by improving mitochondrial enzymes activity and reducing secondary energy loss.

Objective:To investigate the effects of Mor-platin, a novel mitochondrial platinum complex, on proliferation and migration of human hepatoma carcinoma HepG2 cells. Methods:Cell counting kit-8 (CCK-8) assay was used to analyze cell proliferation of Mor-platin and classic anticancer drugs, particularly cisplatin, in HepG2 cells. A laser confocal microscope was used to observe whether Mor-platin can target mitochondria. The morphological changes in cellular mitochondria after treatment with Mor-platin were ob-served on a transmission electron microscope. Cell apoptosis was measured by flow cytometry, and cell invasion was evaluated by three-dimensional tumor spheroid model. Results:Mor-platin can inhibit cell proliferation and is dose dependent. The half inhibitory concentration (IC50) of Mor-platin is lower than that of cisplatin. Laser confocal images showed that Mor-platin can target cell mito-chondria and enrich cell mitochondria. Transmission electron microscopy images showed that cell mitochondrial morphology changed after Mor-platin treatment. Furthermore, cell mitochondrial membrane is incomplete and mitochondrial cristae are reduced. Cell apoptosis caused by Mor-platin is dose dependent. The three-dimensional tumor spheroid model showed that the cell areas of the group subjected to Mor-platin treatment are smaller than those of the control group. Conclusion:Mor-platin can target cell mitochon-dria, change the cell mitochondrial morphology, inhibit cell proliferation, and thus promote cell apoptosis. It also showed better anti-cancer effects than cisplatin. Furthermore, Mor-platin can inhibit three-dimensional tumor spheroid invasion. These results suggest that Mor-platin is a potential antitumor drug.