AIM: To evaluate the effects of survivin on cell proliferation and apoptosis in liver cancer. METHODS: MTT assay was used to generate and optimize phosphorothioate antisense oligonucleotides (ODNs)-LipofectamineTM2000...AIM: To evaluate the effects of survivin on cell proliferation and apoptosis in liver cancer. METHODS: MTT assay was used to generate and optimize phosphorothioate antisense oligonucleotides (ODNs)-LipofectamineTM2000 (LiP) compound by varying ODNs (μg): LiP (μL) ratios from 1:0.5 to 1:5. Then, liver cancer cells (HepG2) were transfected with the compound. By using RT-PCR and Western blot, the expression levels of survivin mRNA and proteins were detected in HepG2 cells treated with antisense compounds (ODNs:LiP=1:4), and compared with those treated with sense compounds (1:4) as control. MTT assay was applied to the determination of cell proliferation in HepG2 cells. Active caspase-3 was evaluated by flow cytometric analysis. The morphological changes were assessed by electron microscopy. Laser scanning confocal microscopy was performed to detect the subcellular localization of survivin proteins in treated and untreated cells. RESULTS: Antisense compounds (1:4) down-regulated survivin expression (mRNA and protein) in a dose-dependent manner with an IC50 of 250 nmol/L. Its maximum effect was achieved at a concentration of 500 nmol/L, at which mRNA and protein levels were down-regulated by 80%. The similar results were found in MTT assay. Antisense compound (l:4)-treated cells revealed increased caspase-3-like protease activity compared with untreated cells. Untreated cells as control were primarily negative for the presence of active-caspase-3. As shown by transmission electron microscopy, treated cells with antisense compounds (1:4) resulted in morphological changes such as blebbing and loss of microvilli, vacuolization in the cytoplasm, condensation of the cytoplasm and nuclei, and fragmented chromatin. Immunofluorescence analysis confirmed the presence of survivin protein pool inside the cytoplasm in untreated cells. Labeled-FITC immunofluorescence staining of survivin clearly showed that survivin was distributed mainly in a spotted form inside the cytoplasm. Whereas cells treated with antisense compounds were rare and weak inside the cytoplasm. CONCLUSION: Down-regulation of survivin expression induced by the antisense compounds reduces tumor growth potential, promotes apoptosis and affects the localization of survivin proteins in HepG2 cells. Furthermore, survivin protein is a key molecule associated with proliferation and apoptosis, and antisense oligonucleotides targeting survivin have a bright prospect in the therapy of liver cancer.展开更多
AIM： To determine which treatment modality - hepatectomy or percutaneous ablation - is more beneficial for patients with small hepatocellular carcinoma （HCC） （≤4 cm） in terms of long-term outcomes. METHODS： A r...AIM： To determine which treatment modality - hepatectomy or percutaneous ablation - is more beneficial for patients with small hepatocellular carcinoma （HCC） （≤4 cm） in terms of long-term outcomes. METHODS： A retrospective analysis of 149 patients with HCC ≤ 4 cm was conducted. Eighty-five patients underwent partial hepatectomy （anatomic in 47 and nonanatomic in 38） and 64 underwent percutaneous ablation （percutaneous ethanol injection in 37, radiofrequency ablation in 21, and microwave coagulation in 6）. The median follow-up period was 69 mo. RESULTS： Hepatectomy was associated with larger tumor size （P〈0.001）, whereas percutaneous ablation was significantly associated with impaired hepatic functional reserve. Local recurrence was less frequent following hepatectomy （P〈0.0001）. Survival was better following hepatectomy （median survival time： 122 mo） than following percutaneous ablation （median survival time： 66 mo; P= 0.0123）. When tumor size was divided into ≤ 2 cm vs 〉 2 cm, the favorable effects of hepatectomy on long-term survival was seen only in patients with tumors 〉2 cm （P= 0.0001）. The Cox proportional hazards regression model revealed that hepatoctomy （P= 0.006） and tumors ≤ 2 cm （P=0.017） were independently associated with better survival. CONCLUSION： Hepatectomy provides both better local control and better long-term survival for patients with HCC ≤4 cm compared with percutaneous ablation. Of the patients with HCC ≤4 cm, those with tumors 〉 2 cm are good candidates for hepatectomy, provided that the hepatic functional reserve of the patient permits resection.展开更多
AIM： To evaluate whether treatment with the Prometheus system significantly affects cytokines, coagulation factors and other plasma proteins. METHODS： We studied nine patients with acute-onchronic liver failure and ...AIM： To evaluate whether treatment with the Prometheus system significantly affects cytokines, coagulation factors and other plasma proteins. METHODS： We studied nine patients with acute-onchronic liver failure and accompanying renal failure. Prometheus therapy was performed on 2 consecutive days for up to 6 h in all patients. Several biochemical parameters and blood counts were assessed at regular time points during Prometheus treatment. RESULTS： We observed a significant decrease of both protein-bound （e.g. bile acids） and water-soluble （e.g. ammonia） substances after Prometheus therapy. Even though leukocytes increased during treatment （P〈 0.01）, we found no significant changes of C-reactive protein, interleukin-6, and tumor necrosis factor-o plasma levels （all P 〉 0.5）. Further, antithrombin 3, factor II and factor V plasma levels did not decrease during Prometheus therapy （all P 〉0.5）, and the INR remained unchanged （P = 0.4）. Plasma levels of total protein, albumin, and fibrinogen were also not altered during Prometheus treatment （all P 〉 0.5）. Finally, platelet count did not change significantly during therapy （P= 0.6）. CONCLUSION： Despite significant removal of protein- bound and water-soluble substances, Prometheus therapy did not affect the level of cytokines, coagulation factors or other plasma proteins. Thus, the filters and adsorbers used in the system are highly effective and specific for water-soluble substances and toxins bound to the albumin fraction.展开更多
AIM: To investigate whether the changes of gap junction gene connexin messenger RNA in the noncancerous liver tissue of patients with hepatocellular carcinoma (HCC) could play a significant role in its postresection r...AIM: To investigate whether the changes of gap junction gene connexin messenger RNA in the noncancerous liver tissue of patients with hepatocellular carcinoma (HCC) could play a significant role in its postresection recurrence.METHODS: Seventy-nine consecutive patients having undergone curative resection for HCC entered this study.Using a reverse-transcription polymerase chain reaction (RT-PCR)-based assay, connexin (Cx) 26, connexin (Cx)32 and connexin (Cx) 43 mRNAs were determined prospectively in noncancerous liver tissues from these 79 patients and in the liver tissues from 15 controls. The correlations between connexin mRNA expression and the clinicopathological variables and outcomes (tumor recurrence and recurrence related mortality) were studied.RESULTS: Compared with liver tissues of control patients,the expression of Cx 32 mRNA in noncancerous liver tissues was significantly lower (mean: 0.715 vscontrol 1.225,P＜0.01), whereas the decreased Cx 26 mRNA (mean:0.700 vs of control 1.205,P＞0.05) and increased Cx 43 mRNA (mean: 0.241 vscontrol 0.100, P＞0.05) had no statistical significance. We defined the value of Cx 32 mRNA or Cx 26mRNA below 0.800 as a lower value. By multivariate analysis for noncancerous livers, a lower value of Cx 32 mRNA correlated significantly with a risk of HCC recurrence and recurrence-related mortality. The lower value of Cx 26 mRNA did not correlate with recurrence and mortality. The increased value of Cx43 mRNA also did not correlate with postoperative recurrence and recurrence-related mortality. By multivariate analysis, other significant predictors of HCC recurrence included vascular permeation, cellular dedifferentiation, and less encaps-ulation. The other significant parameter of recurrence related mortality was vascular permeation.CONCLUSION: The decreased expression of Cx 32 mRNA in noncancerous liver tissues plays a significant role in the prediction of postoperative recurrence of HCC.展开更多
Metastatic melanoma is also a challenge for surgeons. Recently, it has been reported that aggressive surgery combined with supportive therapy may be potential benefit for the condition. Therefore, we report a case of ...Metastatic melanoma is also a challenge for surgeons. Recently, it has been reported that aggressive surgery combined with supportive therapy may be potential benefit for the condition. Therefore, we report a case of ocular melanoma metastatic to multiple visceral sites treated by cytoreductive surgery after initial intra-,arterial hepatic chemoembolization展开更多
AIM: To study genetic difference of mitochondrial DMA (mtDNA) between two hepatocarcinoma cell lines (Hca-F and Hca-P) with diverse metastatic characteristics and the relationship between mtDNA changes in cancer cells...AIM: To study genetic difference of mitochondrial DMA (mtDNA) between two hepatocarcinoma cell lines (Hca-F and Hca-P) with diverse metastatic characteristics and the relationship between mtDNA changes in cancer cells and ttieir oncogenic phenotype. METHODS: Mitochondrial DMA D-loop, tRNAMet+Glu+Ile and ND3 gene fragments from the hepatocarcinoma cell lines with 1100,1126 and 534 bp in length respectively were analysed by PCR amplification and restriction fragment length polymorphism techniques. The D-loop 3′ end sequence of the hepatocarcinoma cell lines was determined by sequencing. RESULTS: No amplification fragment length polymorphism and restriction fragment length polymorphism were observed in tRNAMet+Glu+Ile, ND3 and D-loop of mitochondrial DNA of the hepatocarcinoma cells. Sequence differences between Hca-F and Hca-P were found in mtDNA D-loop. CONCLUSION: Deletion mutations of mitochondrial DNA restriction fragment may not play a significant role in carcinogenesis. Genetic difference of mtDNA D-loop between Hca-F and Hca-P, which may reflect the environmental and genetic influences during tumor progression, could be linked to their tumorigenic phenotypes.展开更多
Objective To observe in vitro effects and morphological changes of human peripheral blood dendritic cells (DCs) on the ability of lymphokine and phytohaemagglutininum (PHA) activated killer (LPAK) cells to induce apo...Objective To observe in vitro effects and morphological changes of human peripheral blood dendritic cells (DCs) on the ability of lymphokine and phytohaemagglutininum (PHA) activated killer (LPAK) cells to induce apoptosis of the human hepatoma cell line (BEL-7402， B).Methods Experimental groups were divided into LD group (DCs+L+B), L group (L+B)， D group (DCs+B) and B group. The methods of neutral red uptake, ordinary light microscopy, electron microscopy, TDT mediated X-dUTP nick end labeling (TUNEL) were used. Results The difference between the D group and the B group was not distinct (P>0.05). The difference between the LD group and the L group was distinct， with DCs+LPAK >LPAK (P<0.01) in cytotoxity. Apoptotic cells were TUNEL positive in light microscopy, and apoptotic nuclei were stained yellow brown and dark brown, with size and shape varying from cell to cell. Ultrastructural change in apoptotic tumor cells comprised of compaction and condensation of nuclear chromatin， and condensation of cytoplasm and apoptotic bodies. At the same time, LPAK cells manifested the characteristics of autophagic apoptosis, and there were some autophagic bodies in it. Conclusions The combination of human blood DCs and LPAK cells could induce apoptosis of BEL-7402 cells effectively, with some LPAK cells manifesting the characteristics of autophagic apoptosis.展开更多
基金Supported by the National Natural Science Foundation of China, No.30171059
文摘AIM: To evaluate the effects of survivin on cell proliferation and apoptosis in liver cancer. METHODS: MTT assay was used to generate and optimize phosphorothioate antisense oligonucleotides (ODNs)-LipofectamineTM2000 (LiP) compound by varying ODNs (μg): LiP (μL) ratios from 1:0.5 to 1:5. Then, liver cancer cells (HepG2) were transfected with the compound. By using RT-PCR and Western blot, the expression levels of survivin mRNA and proteins were detected in HepG2 cells treated with antisense compounds (ODNs:LiP=1:4), and compared with those treated with sense compounds (1:4) as control. MTT assay was applied to the determination of cell proliferation in HepG2 cells. Active caspase-3 was evaluated by flow cytometric analysis. The morphological changes were assessed by electron microscopy. Laser scanning confocal microscopy was performed to detect the subcellular localization of survivin proteins in treated and untreated cells. RESULTS: Antisense compounds (1:4) down-regulated survivin expression (mRNA and protein) in a dose-dependent manner with an IC50 of 250 nmol/L. Its maximum effect was achieved at a concentration of 500 nmol/L, at which mRNA and protein levels were down-regulated by 80%. The similar results were found in MTT assay. Antisense compound (l:4)-treated cells revealed increased caspase-3-like protease activity compared with untreated cells. Untreated cells as control were primarily negative for the presence of active-caspase-3. As shown by transmission electron microscopy, treated cells with antisense compounds (1:4) resulted in morphological changes such as blebbing and loss of microvilli, vacuolization in the cytoplasm, condensation of the cytoplasm and nuclei, and fragmented chromatin. Immunofluorescence analysis confirmed the presence of survivin protein pool inside the cytoplasm in untreated cells. Labeled-FITC immunofluorescence staining of survivin clearly showed that survivin was distributed mainly in a spotted form inside the cytoplasm. Whereas cells treated with antisense compounds were rare and weak inside the cytoplasm. CONCLUSION: Down-regulation of survivin expression induced by the antisense compounds reduces tumor growth potential, promotes apoptosis and affects the localization of survivin proteins in HepG2 cells. Furthermore, survivin protein is a key molecule associated with proliferation and apoptosis, and antisense oligonucleotides targeting survivin have a bright prospect in the therapy of liver cancer.
文摘AIM： To determine which treatment modality - hepatectomy or percutaneous ablation - is more beneficial for patients with small hepatocellular carcinoma （HCC） （≤4 cm） in terms of long-term outcomes. METHODS： A retrospective analysis of 149 patients with HCC ≤ 4 cm was conducted. Eighty-five patients underwent partial hepatectomy （anatomic in 47 and nonanatomic in 38） and 64 underwent percutaneous ablation （percutaneous ethanol injection in 37, radiofrequency ablation in 21, and microwave coagulation in 6）. The median follow-up period was 69 mo. RESULTS： Hepatectomy was associated with larger tumor size （P〈0.001）, whereas percutaneous ablation was significantly associated with impaired hepatic functional reserve. Local recurrence was less frequent following hepatectomy （P〈0.0001）. Survival was better following hepatectomy （median survival time： 122 mo） than following percutaneous ablation （median survival time： 66 mo; P= 0.0123）. When tumor size was divided into ≤ 2 cm vs 〉 2 cm, the favorable effects of hepatectomy on long-term survival was seen only in patients with tumors 〉2 cm （P= 0.0001）. The Cox proportional hazards regression model revealed that hepatoctomy （P= 0.006） and tumors ≤ 2 cm （P=0.017） were independently associated with better survival. CONCLUSION： Hepatectomy provides both better local control and better long-term survival for patients with HCC ≤4 cm compared with percutaneous ablation. Of the patients with HCC ≤4 cm, those with tumors 〉 2 cm are good candidates for hepatectomy, provided that the hepatic functional reserve of the patient permits resection.
文摘AIM： To evaluate whether treatment with the Prometheus system significantly affects cytokines, coagulation factors and other plasma proteins. METHODS： We studied nine patients with acute-onchronic liver failure and accompanying renal failure. Prometheus therapy was performed on 2 consecutive days for up to 6 h in all patients. Several biochemical parameters and blood counts were assessed at regular time points during Prometheus treatment. RESULTS： We observed a significant decrease of both protein-bound （e.g. bile acids） and water-soluble （e.g. ammonia） substances after Prometheus therapy. Even though leukocytes increased during treatment （P〈 0.01）, we found no significant changes of C-reactive protein, interleukin-6, and tumor necrosis factor-o plasma levels （all P 〉 0.5）. Further, antithrombin 3, factor II and factor V plasma levels did not decrease during Prometheus therapy （all P 〉0.5）, and the INR remained unchanged （P = 0.4）. Plasma levels of total protein, albumin, and fibrinogen were also not altered during Prometheus treatment （all P 〉 0.5）. Finally, platelet count did not change significantly during therapy （P= 0.6）. CONCLUSION： Despite significant removal of protein- bound and water-soluble substances, Prometheus therapy did not affect the level of cytokines, coagulation factors or other plasma proteins. Thus, the filters and adsorbers used in the system are highly effective and specific for water-soluble substances and toxins bound to the albumin fraction.
基金Supported by the Grants From Department of Health, National Science Council, Executive Yuan, Taiwan (NSC-89-2314-B-195-027), China
文摘AIM: To investigate whether the changes of gap junction gene connexin messenger RNA in the noncancerous liver tissue of patients with hepatocellular carcinoma (HCC) could play a significant role in its postresection recurrence.METHODS: Seventy-nine consecutive patients having undergone curative resection for HCC entered this study.Using a reverse-transcription polymerase chain reaction (RT-PCR)-based assay, connexin (Cx) 26, connexin (Cx)32 and connexin (Cx) 43 mRNAs were determined prospectively in noncancerous liver tissues from these 79 patients and in the liver tissues from 15 controls. The correlations between connexin mRNA expression and the clinicopathological variables and outcomes (tumor recurrence and recurrence related mortality) were studied.RESULTS: Compared with liver tissues of control patients,the expression of Cx 32 mRNA in noncancerous liver tissues was significantly lower (mean: 0.715 vscontrol 1.225,P＜0.01), whereas the decreased Cx 26 mRNA (mean:0.700 vs of control 1.205,P＞0.05) and increased Cx 43 mRNA (mean: 0.241 vscontrol 0.100, P＞0.05) had no statistical significance. We defined the value of Cx 32 mRNA or Cx 26mRNA below 0.800 as a lower value. By multivariate analysis for noncancerous livers, a lower value of Cx 32 mRNA correlated significantly with a risk of HCC recurrence and recurrence-related mortality. The lower value of Cx 26 mRNA did not correlate with recurrence and mortality. The increased value of Cx43 mRNA also did not correlate with postoperative recurrence and recurrence-related mortality. By multivariate analysis, other significant predictors of HCC recurrence included vascular permeation, cellular dedifferentiation, and less encaps-ulation. The other significant parameter of recurrence related mortality was vascular permeation.CONCLUSION: The decreased expression of Cx 32 mRNA in noncancerous liver tissues plays a significant role in the prediction of postoperative recurrence of HCC.
文摘Metastatic melanoma is also a challenge for surgeons. Recently, it has been reported that aggressive surgery combined with supportive therapy may be potential benefit for the condition. Therefore, we report a case of ocular melanoma metastatic to multiple visceral sites treated by cytoreductive surgery after initial intra-,arterial hepatic chemoembolization
基金Supported by the National Natural Science Foundation of China, No. 39900173
文摘AIM: To study genetic difference of mitochondrial DMA (mtDNA) between two hepatocarcinoma cell lines (Hca-F and Hca-P) with diverse metastatic characteristics and the relationship between mtDNA changes in cancer cells and ttieir oncogenic phenotype. METHODS: Mitochondrial DMA D-loop, tRNAMet+Glu+Ile and ND3 gene fragments from the hepatocarcinoma cell lines with 1100,1126 and 534 bp in length respectively were analysed by PCR amplification and restriction fragment length polymorphism techniques. The D-loop 3′ end sequence of the hepatocarcinoma cell lines was determined by sequencing. RESULTS: No amplification fragment length polymorphism and restriction fragment length polymorphism were observed in tRNAMet+Glu+Ile, ND3 and D-loop of mitochondrial DNA of the hepatocarcinoma cells. Sequence differences between Hca-F and Hca-P were found in mtDNA D-loop. CONCLUSION: Deletion mutations of mitochondrial DNA restriction fragment may not play a significant role in carcinogenesis. Genetic difference of mtDNA D-loop between Hca-F and Hca-P, which may reflect the environmental and genetic influences during tumor progression, could be linked to their tumorigenic phenotypes.
基金fundingfromtheNaturalScience FoundationofGuangdongProvince (No 1995 0 1)andtheScientific ResearchFoundationoftheRailwayMinistr
文摘Objective To observe in vitro effects and morphological changes of human peripheral blood dendritic cells (DCs) on the ability of lymphokine and phytohaemagglutininum (PHA) activated killer (LPAK) cells to induce apoptosis of the human hepatoma cell line (BEL-7402， B).Methods Experimental groups were divided into LD group (DCs+L+B), L group (L+B)， D group (DCs+B) and B group. The methods of neutral red uptake, ordinary light microscopy, electron microscopy, TDT mediated X-dUTP nick end labeling (TUNEL) were used. Results The difference between the D group and the B group was not distinct (P>0.05). The difference between the LD group and the L group was distinct， with DCs+LPAK >LPAK (P<0.01) in cytotoxity. Apoptotic cells were TUNEL positive in light microscopy, and apoptotic nuclei were stained yellow brown and dark brown, with size and shape varying from cell to cell. Ultrastructural change in apoptotic tumor cells comprised of compaction and condensation of nuclear chromatin， and condensation of cytoplasm and apoptotic bodies. At the same time, LPAK cells manifested the characteristics of autophagic apoptosis, and there were some autophagic bodies in it. Conclusions The combination of human blood DCs and LPAK cells could induce apoptosis of BEL-7402 cells effectively, with some LPAK cells manifesting the characteristics of autophagic apoptosis.
